©2008 F. A.

Davis

Urinalysis
and Body Fluids
©2008 F. A. Davis
©2008 F. A. Davis

Urinalysis
and Body Fluids
Fifth Edition

Susan King Strasinger, DA,

MT(ASCP) Faculty Associate
The University of West Florida
Pensacola, Florida
 Marjorie Schaub Di Lorenzo,
BS, MT(ASCP)SH
Adjunct Instructor
Division of Laboratory Sciences
Clinical Laboratory Science
Program University of Nebraska
Medical Center Omaha, Nebraska
Phlebotomy Program
Coordinator Health
Professions
Nebraska Methodist College
Omaha, Nebraska

©2008 F. A. Davis

F. A. Davis Company
1915 Arch Street
Philadelphia, PA 19103
www.fadavis.com

Copyright © 2008 by F. A. Davis Company

Copyright © 2008 by F. A. Davis Company. All rights reserved. This product is protected by copyright.
No part of it may be reproduced, stored in a retrieval system, or transmitted in any form or by any
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Printed in the United States of America

Last digit indicates print number: 10 9 8 7 6 5 4 3 2 1

Acquisitions Editor: Christa Fratantoro

Manager of Content Development: Deborah Thorp
Manager of Art and Design: Carolyn O’Brien

As new scientific information becomes available through basic and clinical research, recommended
treatments and drug therapies undergo changes. The author(s) and publisher have done everything
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publication. The author(s), edi tors, and publisher are not responsible for errors or omissions or for
consequences from application of the book, and make no warranty, expressed or implied, in regard to
the contents of the book. Any practice described in this book should be applied by the reader in
accordance with professional standards of care used in regard to the unique circumstances that may
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for changes and new information regarding dose and contraindications before administering any drug.
 Caution is especially urged when using new or infrequently ordered drugs.

Library of Congress Cataloging-in-Publication Data

Strasinger, Susan King.
Urinalysis and body fluids / Susan King Strasinger, Marjorie Schaub Di Lorenzo ; photography by Bo
Wang … [et al.] ; illustrations by Sherman Bonomelli. — 5th ed.
p. ; cm.
includes bibliographical references and index.
ISBN 978-0-8036-1697-4 (alk. paper)
1. Urine—Analysis. 2. Body fluids—Analysis. 3. Diagnosis, Laboratory. I. Di Lorenzo, Marjorie
Schaub, 1953- II. Title.
[DNLM: 1. Urinalysis—methods. 2. Body Fluids—chemistry. QY 185 S897u 2008]
RB53.S87 2008
616.07’566—dc22 2007017271

Authorization to photocopy items for internal or personal use, or the internal or personal use of
specific clients, is granted by F. A. Davis Company for users registered with the Copyright Clearance
Center (CCC) Transactional Reporting Service, provided that the fee of $.10 per copy is paid directly
to CCC, 222 Rosewood Drive, Danvers, MA 01923. For those organizations that have been granted
a photocopy license by CCC, a separate system of payment has been arranged. The fee code for
users of the Transactional Reporting Service is: 8036-1698/08 $.10.
©2008 F. A. Davis

To Harry, you will always be my Editor-in-Chief

SKS

To my husband, Scott, and my children,

Michael, Christopher, and Lauren
MSD
©2008 F. A. Davis
©2008 F. A. Davis

Preface

brospinal, seminal, synovial, serous, and amniotic

As will be apparent to the readers, the fifth edition of
Urinal ysis and Body Fluids has been substantially
revised and enhanced. However, the objective of the
text—to provide concise, comprehensive, and carefully
structured instruction in the analysis of nonblood body
fluids—remains the same.
This fifth edition has been redesigned to meet the
changes occurring in both laboratory medicine and
instruc tional methodology.
To meet the expanding technical information
required by students in laboratory medicine, all of the
chapters have been updated. Chapter 1 is devoted to
overall laboratory safety and the precautions relating to
urine and body fluid analysis. Chapter 7 addresses
quality assessment and man agement in the urinalysis
laboratory. Preanalytical, analytical, and postanalytical
factors, procedure manuals, current regu latory issues,
and methods for continuous quality improve ment are
stressed. In Chapter 8 the most frequently encountered
diseases of glomerular, tubular, interstitial, and vascular
origin are related to their associated laboratory tests.
To accommodate advances in laboratory testing of cere
fluids, all of the individual chapters have been
enhanced, and addi tional anatomy and physiology
sections have been added for each of these fluids.
Appendix A provides coverage of the ever-increasing
variety of automated instrumentation avail able to the
urinalysis laboratory. Appendix B discusses the
analysis of bronchioalveolar lavage specimens, an area
of the clinical laboratory that has been expanding in
recent years. This fifth edition has been redesigned to
include exten sive multiple choice questions at the end
of each chapter for student review. In response to
readers’ suggestions, the num ber of color slides has
been significantly increased, and the slides are
included within the text to increase user friendli ness.
The text has been extensively supplemented with
tables, summaries, and procedure boxes, and many
figures are now in full color. Case studies in the
traditional format and clini cal situations relating to
technical considerations are included at the end of the
chapters. Answers to the study questions, case
studies, and clinical situations are also included at the
end of the book. Terms in bold italics appear in the
Glossary; abbreviations in bold are listed in
Abbreviations. Additional support is provided to
 adopting instructors in the form of accompanying
test-generating software, an instructor’s man ual with
criticial thinking exercises for each chapter, and
PowerPoint presentations.
We have given consideration to the suggestions of
our previous readers and believe these valuable
suggestions have enabled us to produce a text to meet
the needs of all users.

Susan King Strasinger

Marjorie Schaub Di Lorenzo

vii
©2008 F. A. Davis
©2008 F. A. Davis

Reviewers

Stephen M. Johnson, MS,

MT(ASCP) Program Director
Ellen P. Digan, MA, MT(ASCP) Medical Technology
Professor of Biology Saint Vincent Health Center
Coordinator of Medical Laboratory Technology Erie, Pennsylvania
Program Manchester Community Tech College
Department of Math, Science, and Health Rhoda S. Jost, MSH, MT(ASCP)
Careers Manchester, Connecticut Faculty Program Director
Medical Laboratory Technology
Brenda L. M. Franks, MT(ASCP) Florida Community College at
Point of Care Testing Coordinator Jacksonville Jacksonville, Florida
Methodist Hospital Pathology
Omaha, Nebraska Pam Kieffer, MS, CLS(MCA),
 MT(ASCP) Program Director
Clinical Laboratory Science Amber G.Tuten, MEd, CLDir(NCA),
Rapid City Regional Hospital MT(ASCP) Program Director
Rapid City, South Dakota Medical Laboratory Technology
Cynthia A. Martine, MEd, Okefenokee Technical College
MT(ASCP) Assistant Professor Waycross, Georgia
Department of Clinical Laboratory Sciences
University of Texas Medical Branch
School of Allied Health
Galveston, Texas

Ulrike Otten, MT(ASCP)SC

University of Nebraska Medical Center
Division of Laboratory Sciences
Clinical Laboratory Science Program
Omaha, Nebraska

Kathleen T. Paff, MA, CLS(NCA),

MT(ASCP) Program Director
Medical Laboratory Technology
Kellogg Community College
Battle Creek, Michigan

Kristy Shanahan, MS, NCA,

MT(ASCP) Associate Professor
Medical Laboratory Technology
Oakton Community College
Des Plaines, Illinois ix
©2008 F. A. Davis
©2008 F. A. Davis

Acknowledgments
Many people deserve credit for the help and
encouragement they have provided us in the
preparation of this fifth edition. Our continued
appreciation is also extended to all of the peo ple who
have been instrumental in the preparation of previ ous
editions.
The valuable suggestions from previous readers
and the support from our colleagues at The University
of West Florida, Northern Virginia Community College,
The Univer sity of Nebraska Medical Center, Methodist
Hospital, and Creighton University Medical Center
have been a great asset to us in the production of this
new edition. We thank each and every one of you.
We extend special thanks to the individuals who
have provided us with so many beautiful photographs
for the text over the years: Bo Wang, MD; Donna L.
Canterbury, BA, MT(ASCP)SH; Joanne M. Davis, BS,
MT(ASCP)SH; M. Paula Neumann, MD; Gregory J.
Swedo, MD; and Scott Di Lorenzo, DDS. We also
thank Sherman Bonomelli, MS, for contribut ing original
visual concepts that became the foundation for many of
the line illustrations.
We also appreciate the help and understanding of
our editors at F. A. Davis, Christa Fratantoro, Elizabeth
Zygarewicz, and Deborah Thorp.
 xi
©2008 F. A. Davis
©2008 F. A. Davis

Contents

Sharp Hazards, 5
Chemical Hazards, 6
Chapter 1. Safety in the Chemical Spills, 6
Clinical Laboratory, 1 Chemical Handling, 6
Biological Hazards, 2 Chemical Hygiene Plan, 6
Personal Protective Equipment, 4 Chemical Labeling, 6
Handwashing, 4 Material Data Safety Sheets, 7
Disposal of Biological Waste, 5
Radioactive Hazards, 7
 Electrical Hazards, 7
Fire/Explosive Hazards, 8
Physical Hazards, 8
Chapter 2. Renal Function,
11 Renal Physiology, 12
Renal Blood Flow, 12
Glomerular Filtration, 13
Tubular Reabsorption, 14
Tubular Secretion, 17
Renal Function Tests, 18
Glomerular Filtration Tests, 18
Tubular Reabsorption Tests, 22
Tubular Secretion and
Renal Blood Flow Tests, 24
Chapter 3. Introduction
to Urinalysis, 29
History and Importance, 29
Urine Formation, 31
Urine Composition, 31
Urine Volume, 31
Specimen Collection, 32
Specimen Handling, 33
Specimen Integrity, 33
Specimen Preservation, 33
Types of Specimens, 34
Random Specimen, 34
First Morning Specimen, 34
Fasting Specimen (Second Morning), 34
2-Hour Postprandial Specimen, 35
Glucose Tolerance Specimens, 35
24-Hour (or Timed) Specimen, 36
Catheterized Specimen, 36
Midstream Clean-Catch Specimen, 36
Suprapubic Aspiration, 36
Prostatitis Specimen, 36
Pediatric Specimen, 37
Drug Specimen Collection, 37
Chapter 4. Physical
Examination of Urine, 41
Color, 42
Normal Urine Color, 42
Abnormal Urine Color, 43
Clarity, 44
Normal Clarity, 44
Nonpathologic Turbidity, 44
Pathologic Turbidity, 45
Specific Gravity, 45
Urinometer, 46
Refractometer, 47
Harmonic Oscillation Densitometry, 48
Clinical Correlations, 48
Odor, 49
xiii
©2008 F. A. Davis
xiv CONTENTS
Chapter 5. Chemical
Examination of Urine, 53
Reagent Strips, 54
Reagent Strip Technique, 54
Handling and Storage of Reagent Strips, 55
Quality Control of Reagent Strips, 55
pH, 56
Clinical Significance, 56
Reagent Strip Reactions, 56
Protein, 57
Clinical Significance, 57
Prerenal Proteinuria, 57
Renal Proteinuria, 58
Postrenal Proteinuria, 58
Reagent Strip Reactions, 58
Reaction Interference, 59
Glucose, 61
Clinical Significance, 62
Reagent Strip (Glucose
Oxidase) Reactions, 62
Reaction Interference, 63
Copper Reduction Test, 63
Comparison of Glucose
Oxidase and Clinitest, 64
Ketones, 64
Clinical Significance, 64
Reagent Strip Reactions, 65
Reaction Interference, 65
Blood, 65
 Clinical Significance, 66 Centrifugation, 83
Hematuria, 66 Sediment Preparation, 83
Hemoglobinuria, 66 Volume of Sediment Examined, 83
Myoglobinuria, 66
Examination of the Sediment, 83
Hemoglobinuria Versus Myoglobinuria, 67
Reporting the Microscopic Examination, 84
Reagent Strip Reactions, 67
Correlation of Results, 84
Reaction Interference, 67
Sediment Examination Techniques,
Bilirubin, 68
84 Sediment Stains, 85
Production of Bilirubin, 68
©2008 F. A. Davis
Clinical Significance, 68

Cytodiagnostic Urine Testing, 87

Reagent Strip (Diazo) Reactions, 68 Microscopy, 87
Ictotest Tablets, 68 Types of Microscopy, 89
Reaction Interference, 69 Sediment Constituents, 92
Urobilinogen, 69 Red Blood Cells, 92
Clinical Significance, 70 White Blood Cells, 94
Reagent Strip Reactions and Interference, 70 Epithelial Cells, 95
Reaction Interference, 70 Bacteria, 100
Ehrlich Tube Test, 70 Yeast, 100
Watson-Schwartz Differentiation Test, 71 Parasites, 100
Hoesch Screening Test Spermatozoa, 100
for Porphobilinogen, 71
Mucus, 102
Nitrite, 72 Casts, 102
Clinical Significance, 72 Urinary Crystals, 110
Reagent Strip Reactions, 72 Urinary Sediment Artifacts, 119
Reaction Interference, 73
Leukocyte Esterase, 73 Chapter 7. Quality Assessment
and Management in the
Clinical Significance, 73
Urinalysis Laboratory, 127
Reagent Strip Reaction, 74
Urinalysis Procedure Manual, 128
Reaction Interference, 74
Preanalytical Factors, 129
Specific Gravity, 74
Analytical Factors, 129
Reagent Strip Reaction, 75
Postanalytical Factors, 134
Reaction Interference, 75
Regulatory Issues, 135
Chapter 6. Microscopic Quality Management, 137
Examination of Urine, 81 Medical Errors, 139
Macroscopic Screening, 82
Preparation and Examination Chapter 8. Renal Disease, 143
of the Urine Sediment, 82 Glomerular Disorders, 144
Commercial Systems, 82 Glomerulonephritis, 144
Specimen Preparation, 83 Acute Poststreptococcal Glomerulonephritis, 144
Specimen Volume, 83 Rapidly Progressive (Crescentic)
 Glomerulonephritis, 144 Carbohydrate Disorders, 170
Goodpasture Syndrome, 144
Membranous Glomerulonephritis, 145 Chapter 10. Cerebrospinal Fluid,
Membranoproliferative 177 Formation and Physiology, 178
Glomerulonephritis, 145 Specimen Collection
Chronic Glomerulonephritis, 145 and Handling, 178
Immunogloblin A Nephropathy, 145 Appearance, 179
Nephrotic Syndrome, 145 Traumatic Collection (Tap), 179
©2008 F. A. Davis

CONTENTS xvxvi CONTENTS

Minimal Change Disease, 146 Uneven Distribution of Blood, 179

Focal Segmental Glomerulosclerosis, Clot Formation, 180
146 Alport Syndrome, 147 Xanthochromic Supernatant, 180
Diabetic Nephropathy, 147 Cell Count, 180
Tubular Disorders, 147 Methodology, 181

Acute Tubular Necrosis, 149 Total Cell Count, 181

WBC Count, 181
Hereditary and Metabolic
Tubular Disorders, 149 Corrections for Contamination, 182
Fanconi Syndrome, 149 Quality Control of Cerebrospinal Fluid and Other Body Fluid
Cell Counts, 182
Nephrogenic Diabetes Insipidus, 149
Differential Count on a Cerebrospinal Fluid
Renal Glycosuria, 149
Specimen, 182
Interstitial Disorders, 149
Cytocentrifugation, 182
Acute Pyelonephritis, 150
Cerebrospinal Fluid Cellular Constituents,
Chronic Pyelonephritis, 150 183
Acute Interstitial Nephritis, 151 Chemistry Tests, 189
Renal Failure, 151 Cerebrospinal Protein, 189
Renal Lithiasis, 152 Cerebrospinal Fluid Glucose, 191
Cerebrospinal Fluid Lactate, 192
Chapter 9. Urine Screening Cerebrospinal Fluid Glutamine, 192
for Metabolic Disorders,
Microbiology Tests, 192
159
Gram Stain, 193
Overflow Versus
Renal Disorders, 160 Serologic Testing, 194
Newborn Screening Tests, 160 Teaching Cerebrospinal
Fluid Analysis, 195
Amino Acid Disorders, 161
Phenylalanine-Tyrosine Disorders, 161 Chapter 11. Semen, 199
Branched-Chain Amino Acid Disorders,
Physiology, 199
164 Tryptophan Disorders, 165
Specimen Collection, 200
Cystine Disorders, 166
Specimen Handling, 201
Porphyrin Disorders, 167 Semen Analysis, 201
Historical Note, 168 Appearance, 201
Mucopolysaccharide Disorders, Liquefaction, 201
169 Purine Disorders, 170 Volume, 201
Viscosity, 202 228
pH, 202 Appearance, 228
Sperm Concentration/Count, 202 Laboratory Tests, 228
©2008 F. A. Davis
Sperm Motility, 203
Sperm Morphology, 203

Peritoneal Fluid, 229

Transudates Versus Exudates, 229
Additional Testing, 205 Appearance, 230
Sperm Viability, 205 Laboratory Tests, 230
Seminal Fluid Fructose, 206
Antisperm Antibodies, 206 Chapter 14. Amniotic Fluid,
Microbial and Chemical Testing, 207 235 Physiology, 235
Postvasectomy Semen Analysis, 207 Function, 235
Sperm Function Tests, 207 Volume, 236
Semen Analysis Quality Control, 207 Chemical Composition, 236
Differentiating Maternal Urine
Chapter 12. Synovial Fluid, 211 From Amniotic Fluid, 237
Physiology, 211 Specimen Collection, 237
Specimen Collection Indications for Amniocentesis, 237
and Handling, 212 Indications for Performing Amniocentesis,
Color and Clarity, 213 237 Collection, 237
Viscosity, 213
Specimen Handling
Cell Counts, 213 and Processing, 237
Differential Count, 213 Color and Appearance, 238
Crystal Identification, 214
Tests for Fetal Distress, 238
Types of Crystals, 214
Hemolytic Disease of the Newborn, 238
Slide Preparation, 215
Neural Tube Defects, 239
Crystal Polarization, 216
Tests for Fetal Maturity, 239
Chemistry Tests, 217
Fetal Lung Maturity, 239
Microbiologic Tests, 217
Lecithin-Sphingomyelin Ratio, 240
Serologic Tests, 218
Amniostat-FLM, 240
Chapter 13. Serous Fluid, 221 Foam Stability, 240
Formation, 221 Microviscosity: Fluorescence
Specimen Collection Polarization Assay, 241
and Handling, 222 Lamellar Bodies and Optical
Density, 241
Transudates and Exudates, 223 General
Laboratory Procedures, 223 Pleural Fluid,
223 CONTENTS xvii

Appearance, 224 Chapter 15. Fecal Analysis,

Hematology Tests, 225
245 Physiology, 245
Chemistry Tests, 226
Diarrhea, 246
Microbiologic and Serologic Tests, 227 Pericardial Fluid,
Steatorrhea, 248
 252 Fecal Enzymes, 253
Specimen Collection, 248
Carbohydrates, 253
Macroscopic Screening, 248
Color, 248 Appendix A, 259
Appearance, 248
Appendix B, 265
Microscopic Examination
of Feces, 249 Answers to Case Studies
Fecal Leukocytes, 249 and Clinical Situations, 267
Muscle Fibers, 249
Answers to Study Questions,
Qualitative Fecal Fats, 249
Chemical Testing of Feces, 250 273 Abbreviations, 277
Occult Blood, 250 Glossary, 279
Quantitative Fecal Fat Testing, 251
APT Test (Fetal Hemoglobin), Index, 285
©2008 F. A. Davis
©2008 F. A. Davis

1 R

Safety in the Clinical

Laboratory
LEARNING OBJECTIVES
Upon completion of this chapter, the reader will be able to:
used.
1 List the components of the chain of infection
and the laboratory safety precautions that 5 Correctly perform routine handwashing. 6
break the chain. Describe the acceptable methods for
disposing of biological waste and sharp
2 Differentiate among and state the
precautions addressed by Universal objects in the uri nalysis laboratory.
Precautions, body sub stance isolation, and
Standard Precautions.
3 State the specifics of the Occupational KEY TERMS
Exposure to Blood-Borne Pathogens 7 Discuss the components and purpose of
Standard. chemical hygiene plans and material safety data
4 Describe the types of personal protective sheets.
equip ment that laboratory personnel wear, 8 State the components of the National Fire
including when, how, and why each article is
 Protec tion Association hazardous material taken when a fire is discovered.
labeling system. 11 Differentiate among class A, B, C, and D
9 Describe precautions that laboratory fires with regard to material involved and
personnel should take with regard to methods of extinguishing each type.
radioactive and elec trical hazards. 12 Recognize standard hazard warning symbols.
10 Explain the RACE and PASS actions to be
Occupational Safety (PPE)
and Health postexposure
Administration prophylaxis (PEP)
(OSHA) radioisotope
biohazardous
personal Standard Precautions
chain of infection
protective Universal Precautions
chemical hygiene plan
equipment (UP)
Material Safety Data Sheet

1
©2008 F. A. Davis

2 CHAPTER 1 • Safety in the Clinical Laboratory

Table 1–1 Types of Safety Hazards

Type Source Possible Injury
infections Cuts, punctures, or
Biological Preservatives and reagents
blood-borne pathogen exposure
Sharps Equipment and radioisotopes
Exposure to toxic, carcinogenic, or
Chemical Ungrounded or wet equipment;
caustic agents Radiation exposure
Radioactive Electrical frayed cords Bunsen burners,
Fire/explosive Physical organic chemicals Wet floors, heavy Burns or shock
boxes, patients Burns or dismemberment
Infectious agents
Bacterial, fungal, viral, or parasitic Falls, sprains, or strains
Needles, lancets, broken glass

From Strasinger, SK and DiLorenzo, MA: Phlebotomy Workbook for the Multiskilled Healthcare Professional, FA Davis,
Philadelphia, 1996, p 62, with permission.

■ ■ ● Biological Hazards
The clinical laboratory contains a variety of safety The health-care setting provides abundant
hazards, many of which are capable of producing
sources of potentially harmful microorganisms.
serious injury or life threatening disease. To work safely
These microor ganisms are frequently present
in this environment, labo ratory personnel must learn
in the specimens received in the clinical
what hazards exist, the basic safety precautions
laboratory. Understanding
associated with them, and how to apply the basic rules
how microorganisms are transmitted (chain of
of common sense required for everyday safety. As can
infection) is essential to preventing infection. The
be seen in Table 1–1, some hazards are unique to the
chain of infection requires a continuous link between a
health-care environment, and others are encountered
source, a method of transmission, and a susceptible
rou tinely throughout life.
host. The source is the location of potentially harmful
microorganisms, such as a contami nated clinical
 specimen or an infected patient. Microorganisms of infection is complete, the infected host then
becomes another source able to transmit the
microorganisms to others.
In the clinical laboratory, the most direct contact
Hand washing
with a source of infection is through contact with patient
Biohazardous waste disposal
Decontamination specimens, although contact with patients and infected
Specimen bagging objects also occurs. Preventing completion of the chain
of infection is a primary objective of biological safety.
SOURCE Figure 1-1 uses the universal symbol for biohazardous
material to demonstrate how following prescribed
OI N
S
safety practices can break the chain of infection. Figure
H
1-1 places particular emphasis on labora tory practices.
from the source are transmitted to the host. This may
Proper handwashing and wearing personal
occur by direct contact (e.g., the host touches the
protective equipment (PPE) are of major importance
patient, specimen, or a contaminated object), inhalation
in the laboratory. Concern over exposure to
of infected material (e.g., aerosol droplets from a
blood-borne pathogens, primarily hepatitis B virus
patient or an uncapped centrifuge tube), ingestion of a
(HBV) and human immunodeficiency virus (HIV),
contaminated substance (e.g., food, water, specimen),
resulted in the drafting of guidelines and regulations
or from an animal or insect vector bite. Once the chain
Immunization practices related to the
Healthy lifestyle
biohazard symbol.
Exposure control plan
S
O
(Adapted from
Strasinger, SK and
T
S
I

TR
AN
S M DiLorenzo, MA:
Hand washing Phlebotomy Work book
Personal protective for the Multiskilled
equipment Aerosol
prevention
Healthcare Professional,
Sterile/disposable FA
equipment
Figure 1–1 Chain of
infection and safety
Standard precautions
Postexposure prophylaxis Davis, Philadelphia, 1996, p 62, with permission.)
Pest control
©2008 F. A. Davis Standard Precautions are as follows:

1. Handwashing: Wash hands after touching blood,

body fluids, secretions, excretions, and contaminated items,
by the Centers for Disease Control and Prevention (CDC) whether or not gloves are worn. Wash hands
and the Occupational Safety and Health Administration immediately after gloves are removed, between
(OSHA) to prevent exposure. In 1987 the CDC instituted patient contacts, and when otherwise indicated to
Universal Precautions (UP). Under UP all patients are con avoid transfer of microorganisms to other patients
sidered to be possible carriers of blood-borne pathogens. The or environments. Washing hands may be necessary
guideline recommends wearing gloves when collecting or between tasks and procedures on the same patient to
handling blood and body fluids contaminated with blood and prevent cross-contamination of different body sites.
wearing face shields when there is danger of blood splashing 2. Gloves: Wear gloves (clean, nonsterile gloves are
on mucous membranes and when disposing of all needles adequate) when touching blood, body fluids, secre
and sharp objects in puncture-resistant containers. The CDC tions, excretions, and contaminated items. Put on
excluded urine and body fluids not visibly contaminated by gloves just before touching mucous membranes and
blood from UP, although many specimens can contain a con nonintact skin. Change gloves between tasks and
siderable amount of blood before it becomes visible. The procedures on the same patient after contact with
modification of UP for body substance isolation (BSI) material that may contain a high concentration of
helped to alleviate this concern. BSI guidelines are not limited microorganisms. Remove gloves promptly after use,
to blood-borne pathogens; they consider all body fluids and before touching noncontaminated items and environ mental
moist body substances to be potentially infectious. According surfaces, and before going to another patient. Always wash
to BSI guidelines, personnel should wear gloves at all times your hands immediately after glove
when encountering moist body substances. A major disad removal to avoid transfer of microorganisms to other patients
vantage of BSI guidelines are that they do not recommend or environments.
handwashing following removal of gloves unless visual con
tamination is present. 3. Mask, eye protection, and face shield: Wear a mask and
In 1996 the CDC combined the major features of UP and BSI eye protection or a face shield to protect mucous membranes
guidelines and called the new guidelines Standard Pre of the eyes, nose, and mouth during pro cedures and patient
cautions. Although Standard Precautions, as described care activities that are likely to
below, stress patient contact, the principles most certainly generate splashes or sprays of blood, body fluids,
can also be applied to handling patient specimens in the
laboratory.1

CHAPTER 1 • Safety in the Clinical Laboratory 3
secretions, or excretions. A specially fitted respirator (N95)
must be used during patient care activites related to
suspected mycobacterium exposure.
4. Gown: Wear a gown (a clean, nonsterile gown is ade
quate) to protect skin and to prevent soiling of cloth ing during
procedures and patient care activities that are likely to
generate splashes or sprays of blood, body fluids, secretions,
or excretions. Select a gown that is appropriate for the activity
and the amount of fluid likely to be encountered (e.g.,
fluid-resistant in the laboratory). Remove a soiled gown as
promptly as possible, and wash hands to avoid the transfer of
microorganisms to other patients or environments.
5. Patient care equipment: Handle used patient care
equipment soiled with blood, body fluids, secretions, and
excretions in a manner that prevents skin and mucous
membrane exposures, contamination of clothing, and transfer
of microorganisms to other patients or environments. Ensure
that reusable equip ment is not used for the care of another
patient until it has been cleaned and reprocessed
appropriately. Ensure that single-use items are discarded
properly.
6. Environmental control: Ensure that the hospital has
adequate procedures for the routine care, cleaning, and
disinfection of environmental surfaces, beds, bedrails,
bedside equipment, and other frequently touched surfaces.
Ensure that these procedures are being followed.
7. Linen: Handle, transport, and process linen soiled with
blood, body fluids, secretions, and excretions in a manner
that prevents skin and mucous membrane exposures and
contamination of clothing and that avoids the transfer of
microorganisms to other patients and environments.
8. Occupational health and blood-borne pathogens: Take
care to prevent injuries when using needles, scalpels, and
other sharp instruments or devices;
when handling sharp instruments after procedures; when
cleaning used instruments; and when dispos ing of used
needles. Never recap used needles or otherwise manipulate
them using both hands or use any other technique that
involves directing the point of a needle toward any part of the
body; rather, use self-sheathing needles or a mechanical
device to con ceal the needle. Do not remove used needles
from disposable syringes by hand, and do not bend, break, or
otherwise manipulate used needles by hand. Place used
disposable syringes and needles, scalpel blades, and other
sharp items in appropriate puncture-resistant containers,
which are located as close as practical to the area in which
the items were used, and place reusable syringes and
needles in a puncture-resistant container for transport to the
reprocessing area. Use mouthpieces, resuscitation bags, or
other ventilation devices as an alternative
©2008 F. A. Davis
4 CHAPTER 1 • Safety in the Clinical Laboratory
to mouth-to-mouth resuscitation methods in areas
where the need for resuscitation is predictable.
9. Patient placement: Place a patient who
contami nates the environment or who does
not (or cannot
be expected to) assist in maintaining appropriate
hygiene or environment control in a private room.
If a private room is not available, consult with
infec tion control professionals regarding
patient place
ment or other alternatives.
The Occupational Exposure to Blood-Borne
Pathogens Standard is a law monitored and enforced
by OSHA.2 Specific requirements of this OSHA
standard include the following:
1. Requiring all employees to practice
UP/Standard Pre cautions
2. Providing laboratory coats, gowns, face and
respira tory protection, and gloves to
employees and laun
dry facilities for nondisposable protective clothing
3. Providing sharps disposal containers and
prohibiting recapping of needles
4. Prohibiting eating, drinking, smoking, and
applying cosmetics, lip balm, and contact lens in
the work
area
5. Labeling all biohazardous material and
containers 6. Providing free immunization for
HBV
7. Establishing a daily disinfection protocol for
work surfaces; an appropriate disinfectant for
blood-borne pathogens is sodium hypochlorite
(household bleach diluted 1:10)
8. Providing medical follow-up for employees who
have been accidentally exposed to blood-borne
pathogens
9. Documenting regular training in safety
standards for employees
Any accidental exposure to a possible
blood-borne pathogen must be immediately reported.
Evaluation of the incident must begin right away to
ensure appropriate postex posure prophylaxis
(PEP). The CDC provides periodically updated
guidelines for the management of exposures and rec
ommended PEP.3,4
Personal Protective Equipment
PPE used in the laboratory includes gloves,
fluid-resistant gowns, eye and face shields, and
Plexiglas countertop shields. Gloves should be worn
when in contact with patients, speci mens, and
laboratory equipment or fixtures. When specimens are
collected, gloves must be changed between every
patient. In the laboratory, they are changed whenever
they become noticeably contaminated or damaged
and are always removed when leaving the work area.
Wearing gloves is not a substitute for handwashing,
and hands must be washed after gloves are removed.
A variety of gloves are available, including sterile
and nonsterile, powdered and unpowdered, and latex
and nonla tex. Allergy to latex is increasing among
health-care workers,

and laboratory personnel should be alert for symptoms
of reactions associated with latex. Reactions to latex
include irri tant contact dermititis, which produces
patches of dry, itchy irritation on the hands; delayed
hypersensitivity reactions resembling poison ivy that
appear 24 to 48 hours following exposure; and true,
immediate hypersensitivity reactions often
characterized by facial flushing and breathing difficul
ties. Handwashing immediately after removing gloves
and avoiding powdered gloves may aid in preventing
the devel opment of latex allergies. Replacing latex
gloves with nitrile or vinyl gloves provides an
alternative. Any symptoms of latex allergy should be
reported to a supervisor because true latex allergy can
be life-threatening.5
Fluid-resistant laboratory coats with wrist cuffs are
worn to protect clothing and skin from exposure to
patients’ body substances. These coats should always
be completely but toned, and gloves should be pulled
over the cuffs. They are worn at all times when working
with patient specimens and are removed prior to
leaving the work area. They are changed when they
become visibly soiled. Disposable coats are placed in
containers for biohazardous waste, and nondisposable
coats are placed in designated laundry receptacles.
The mucous membranes of the eyes, nose, and
mouth must be protected from specimen splashes and
aerosols. A variety of protective equipment is available,
including gog gles, full-face plastic shields, and
Plexiglas countertop shields. Particular care should be
©2008 F. A. Davis
taken to avoid splashes and aerosols when removing
container tops, pouring specimens, and cen trifuging
specimens. Specimens must never be centrifuged in
uncapped tubes or in uncovered centrifuges. When
speci mens are received in containers with
contaminated exteriors, the exterior of the container
must be disinfected or, if neces sary, a new specimen
may be requested.
Handwashing
Handwashing is emphasized in Figure 1-1 and in the
Stan dard Precautions guidelines. Hand contact is the
primary method of infection transmission. Laboratory
personnel must always wash hands after gloves are
removed, prior to leaving the work area, at any time
when hands have been knowingly contaminated, before
going to designated break areas, and before and after
using bathroom facilities.
Correct handwashing technique is shown in Figure
1-2 and includes the following steps:
1. Wet hands with warm water.
2. Apply antimicrobial soap.
3. Rub to form a lather, create friction, and
loosen debris.
4. Thoroughly clean between fingers, including
thumbs, under fingernails and rings, and up to
the wrist, for at least 15 seconds.
5. Rinse hands in a downward position.
6. Dry with a paper towel.
7. Turn off faucets with a clean paper towel to
prevent recontamination.
CHAPTER 1 • Safety in the Clinical Laboratory 5
 Figure 1–2 Handwashing technique. (A) Wetting hands. (B) Lathering hands and creating friction. (C) Cleaning
between fingers. (D) Rinsing hands. (E) Drying hands. (F) Turning off water. (From Strasinger, SK and DiLorenzo,
MA: Skills for the Patient Care Technician, FA Davis, Philadelphia, 1999, p 70, with permission.)
Absorbent materials used for cleaning countertops and
removing spills must be discarded in bio hazard
Disposal of Biological Waste containers. Empty urine containers can be discarded
as nonbiologically hazardous waste (Fig. 1-3).
All biological waste, except urine, must be placed in
appropri ate containers labeled with the biohazard
symbol (see Fig. 1-1). This includes both specimens ■ ■ ● Sharp Hazards
and the materials with which the specimens come in
Sharp objects in the laboratory, including
contact. The waste is then decontaminated following
needles, lancets, and broken glassware,
institutional policy: incineration, autoclaving, or pickup
present a serious biological hazard,
by a certified hazardous waste company.
particularly for the transmission of blood-borne
Urine may be discarded by pouring it into a
pathogens. All sharp objects must
laboratory sink. Care must be taken to avoid splashing, ©2008 F. A. Davis
and the sink should be flushed with water after
specimens are discarded. Disinfection of the sink using
a 1:5 or 1:10 dilution of sodium hypochlorite should be 6 CHAPTER 1 • Safety in the Clinical Laboratory
performed daily. Sodium hypochlorite dilutions stored in
plastic bottles are effective for 1 month if
protected from light after preparation. The same
solution also can be used for routinely disinfecting
countertops and acci dental spills. The solution should
be allowed to air-dry on the contaminated area.

A B
Figure 1–3 Technologist disposing of urine (A) sample
and (B) container.
be disposed in puncture-resistant containers.
Puncture-resis tant containers should be
conveniently located within the work area.
■ ■ ● Chemical Hazards
The same general rules for handling
biohazardous materials apply to chemically
hazardous materials; that is, to avoid getting
these materials in or on bod ies, clothes, or work
area. Every chemical in the workplace should be
presumed hazardous.
Chemical Spills
When skin contact occurs, the best first aid is to
flush the area with large amounts of water for at
least 15 minutes and then
seek medical attention. For this reason, all
laboratory person nel should know the location and
proper use of emergency showers and eye wash
stations. Contaminated clothing should be removed
as soon as possible. No attempt should be made to
neutralize chemicals that come in contact with the
skin. Chemical spill kits containing protective
apparel, nonre active absorbent material, and bags
for disposal of contami nated materials should be
available for cleaning up spills.
Chemical Handling
Chemicals should never be mixed together unless
specific instructions are followed, and they must be
added in the order specified. This is particularly
important when combin ing acid and water. Acid
should always be added to water to avoid the
possibility of sudden splashing caused by the rapid
generation of heat in some chemical reactions.
Wearing gog gles and preparing reagents under a
fume hood are recom mended safety precautions.
Chemicals should be used from containers that are
of an easily manageable size. Pipetting by mouth is
unacceptable in the laboratory. State and federal
reg ulations are in place for the disposal of
chemicals and should be consulted.
Chemical Hygiene Plan
OSHA also requires all facilities that use hazardous
chemicals to have a written chemical hygiene plan
(CHP) available to employees.6 The purpose of the
plan is to detail the following:
1. Appropriate work practices
2. Standard operating procedures
3. PPE
4. Engineering controls, such as fume hoods
and flam mables safety cabinets
5. Employee training requirements
6. Medical consultation guidelines
Each facility must appoint a chemical hygiene
officer, who is responsible for implementing and
documenting com pliance with the plan. Examples
of required safety equipment and information are
shown in Figure 1-4.
Chemical Labeling
Hazardous chemicals should be labeled with a
description of their particular hazard, such as
poisonous, corrosive, or car cinogenic. The
National Fire Protection Association (NFPA) has
developed the Standard System for the
Identification of the Fire Hazards of Materials, NFPA
704.7 This symbol system is used to inform fire
fighters of the hazards they may encounter with
fires in a particular area. The diamond-shaped,
color-coded symbol contains information relating to
health, flammability, reactivity, and personal
protection/special pre cautions. Each category is
graded on a scale of 0 to 4, based on the extent of
concern. These symbols are placed on doors,
cabinets, and containers. An example of this system
 is shown in Figure 1-5.
©2008 F. A. Davis Material Data Safety Sheets
The OSHA Federal Hazard Communication
Standard requires that all employees have a right
to know about all chemical haz ards present in their
workplace. The information is provided in the form
of Material Safety Data Sheets (MSDSs) on file in
the workplace. By law, vendors are required to
provide these sheets to purchasers; however, the
facility itself is responsible for obtaining and making
MSDSs available to employees. Information
contained in an MSDS includes the following:
1. Physical and chemical characteristics
2. Fire and explosion potential
3. Reactivity potential
4. Health hazards and emergency first aid
procedures 5. Methods for safe handling
and disposal

Figure 1–4 Chemical safety aids. (A) Equipment. (B)
Informa tion and supplies. (From Strasinger, SK and
DiLorenzo, MA: Skills for the Patient Care Technician,
FA Davis, Philadelphia, 1999, p 70, with permission.)
HAZARDOUS MATERIALS
CLASSIFICATION
HEALTH HAZARD FIRE HAZARD
Flash Point
■ ■ ● Radioactive Hazards
Radioactivity is encountered in the clinical
labora tory when procedures using
radioisotopes are per formed. The
amount of radioactivity present in the
clinical laboratory is very small and
represents little
danger; however, the effects of radiation are
cumulative related to the amount of exposure. The
amount of radiation exposure is related to a
combination of time, distance, and shielding.
Persons working in a radioactive environment are
required to wear measuring devices to determine
the amount of radiation they are accumulating.
Laboratory personnel should be familiar with
the radioactive hazard symbol shown here. This
symbol must be displayed on the doors of all areas
where radioactive material is present. Exposure to
radiation during pregnancy presents a danger to
the fetus; personnel who are pregnant or think they
may be should avoid areas with this symbol.

CHAPTER 1 • Safety in the Clinical Laboratory 7

of electrical safety circuits, and report them
4 Deadly observed outside the Oxidizer
3 Extreme Danger 2 Hazardous
1 Slightly Hazardous 0 Normal Material workplace apply. The Acid
danger of water or fluid Alkali

2 SPECIFIC HAZARD
coming in contact with
equipment is greater in the
laboratory setting.
Corrosive
Use No Water Radiation
OXY ACID ALK COR W
4 May deteriorate 3 Shock and heat may
4 Below 73 F 3 Below 100 F 2 Below 200

31W
deteriorate 2 Violent chemical
F 1 Above 200 F 0 Will not burn Equipment should not be change
operated with wet hands. 1 Unstable if
■ ■ ● Electrical Designated hospital heated
0 Stable
Hazards personnel mon itor
electrical equipment to the appropriate persons.
The laboratory setting closely; however, Equipment that has
contains a large amount of laboratory person nel become wet should be
electrical equipment with should continually observe unplugged and allowed to
which workers have fre for any dangerous dry completely before
REACTIVITY conditions, such as frayed reusing. Equipment also
quent contact. The cords and overloaded should be unplugged
same general rules
 before clean ing. All pronged plugs. electrical source must be
electrical equipment must When an accident involving removed immediately. This
be grounded with three electrical shock occurs, the must be
Figure 1–5 NFPA hazardous material symbols. involved
done without touching the person or the equipment
©2008 F. A. Davis

8 CHAPTER 1 • Safety in the Clinical Laboratory

Table 1–2 Types of Fires and Fire

Extinguishers
Type of Fire
Fire Type Extinguishing Material Composition of Fire Extinguisher
chemicals Electrical chemicals, carbon dioxide,
Class A Class B Class C None
Combustible metals Class ABC or halon
Class D
Class A Water
Wood, paper, clothing Sand or dry powder
Class B Dry chemicals, carbon Dry chemicals
Flammable organic
Class C dioxide, foam, or halon Dry

From Strasinger, SK and DiLorenzo, MA: Skills for the Patient Care Technician, FA Davis, Philadelphia, 1999, p 70, with permission.
in order to avoid transference of the current. Turning off
the circuit breaker, unplugging the equipment, or
moving the equipment using a nonconductive glass or
wood object are safe procedures to follow.
■ ■ ● Fire/Explosive Hazards
The Joint Commission on Accreditation of
Health care Organizations (JCAHO) requires
that all health-care institutions post evacuation
routes and detailed plans to follow in the event
of a fire. Labo
ratory personnel should be familiar with these
procedures. When a fire is discovered, all employees
are expected to take the actions in the acronym RACE:
Rescue—rescue anyone in immediate danger
Alarm—activate the institutional fire alarm
system Contain—close all doors to potentially
affected areas
common, but the label should always be checked
before using. It is important to be able to operate the
fire extin guishers. The acronym PASS can be used to
remember the steps in the operation:
1. Pull pin
2. Aim at the base of the fire
3. Squeeze handles
4. Sweep nozzle side to side.
■ ■ ● Physical Hazards
Physical hazards are not unique to the
laboratory, and routine precautions observed
outside the work place apply. General
precautions to consider are to avoid running in
rooms and hallways, watch for wet
floors, bend the knees when lifting heavy objects, keep
long hair pulled back, avoid dangling jewelry, and
maintain a clean, organized work area. Closed-toe
shoes that provide maximum support are essential for
safety and comfort.

Extinguish—attempt to extinguish the fire, if

possible; exit the area
As discussed previously, laboratory workers often
use potentially volatile or explosive chemicals that
require special procedures for handling and storage.
Flammable chemicals should be stored in safety
cabinets and explo sion-proof refrigerators, and
cylinders of compressed gas should be located away
from heat and securely fastened to a stationary device
to prevent accidental capsizing. Fire blan kets may be
present in the laboratory. Persons with burning clothes
should be wrapped in the blanket to smother the
flames.
The NFPA classifies fires with regard to the type of
burn ing material. It also classifies the type of fire
extinguisher that is used to control them. This
information is summarized in Table 1–2. The
multipurpose ABC fire extinguishers are the most
References
1. Centers for Disease Control and Prevention: Guideline
for Isola tion Precautions in Hospitals, Parts I and II. Web
site: http://www.cdc.gov
2. Occupational Exposure to Blood-Borne Pathogens,
Final Rule. Federal Register 29 (Dec 6), 1991.
3. Centers for Disease Control and Prevention. Updated U.S.
Public Health Service Guidelines for the Management of
Occupational Exposures to HBV, HCV and HIV and
Recommendations for Post-exposure Propylaxis. MMWR
June 29, 2001: 50(RR11); 1-42. Web site:
http://www.cdc.gov
4. Centers for Disease Control and Prevention. Updated U.S.
Public Health Service Guidelines for the Management of
Occupational Exposure to HIV and Recommendations for
Post-exposure Pro
phylaxis. MMWR September 17, 2005: 54(RR09); 1-17.
Web site: http://www.cdc.gov
 5. NIOSH Alert. Preventing Allergic Reactions to Natural
Rubber Latex in the Workplace. DHHS (NIOSH)
Publication 97-135. National Institute for Occupational
Safety and Health, Cincin nati, Ohio, 1997.
6. Occupational Exposure to Hazardous Chemicals in
Laboratories, Final Rule. Federal Register 55 (Jan 31),
1990.
7. National Fire Protection Association: Hazardous
Chemical Data, No. 49. Boston, NFPA, 1991.
©2008 F. A. Davis
STUDY
QUESTIONS
1. In the urinalysis laboratory the primary
source in the chain of infection would be:
A. Patients
B. Needlesticks
C. Specimens
D. Biohazardous waste
2. The best way to break the chain of infection is:
A. Handwashing
B. Personal protective equipment
C. Aerosol prevention
D. Decontamination
3. Standard Precautions differ from Universal
Precautions and body substance isolation by
requiring:
A. Wearing face shields and gloves
whenever blood may be encountered
B. Wearing gloves when encountering any
moist body fluid
C. Washing hands after removing gloves if
visual con tamination is present
D. Wearing gloves when exposed to moist
body fluids and washing hands after glove
removal
4. An employee who is accidentally exposed to
a possible blood-borne pathogen should
immediately:
A. Report to a supervisor
B. Flush the area with water
C. Clean the area with disinfectant
D. Receive HIV propylaxis
5. Personnel in the urinalysis laboratory should
wear lab coats that:
A. Do not have buttons
B. Are fluid-resistant
C. Have short sleeves
D. Have full-length zippers
6. All of the following should be discarded in
biohaz ardous waste containers except:
A. Urine specimen containers
B. Towels used for decontamination
C. Disposable lab coats
D. Blood collection tubes
7. An employer who fails to provide sufficient
gloves for the employees may be fined by
the:
A. CDC
B. NFPA
C. OSHA
D. FDA
8. An acceptable disinfectant for blood and
body fluid decontamination is:
A. Sodium hydroxide
B. Antimicrobial soap
C. Hydrogen peroxide
D. Sodium hypochlorite
CHAPTER 1 • Safety in the Clinical Laboratory 9
9. Proper handwashing includes all of the
following except:
A. Using warm water
B. Rubbing to create a lather
C. Rinsing hands in a downward position
D. Turning on the water with a paper towel
10. Centrifuging an uncapped specimen may
produce a biological hazard in the form of:
A. Vectors
B. Sharps contamination
C. Aerosols
D. Specimen contamination
11. An employee who accidently spills acid on
his arm should immediately:
A. Neutralize the acid with a base
B. Hold the arm under running water for 15
minutes C. Consult the MSDSs
D. Wrap the arm in gauze and go to the
emergency room
12. When combining acid and water,
ensure that: A. Acid is added to
water
B. Water is added to acid
C. They are added simultaneously
D. Water is slowly added to acid
13. An employee can learn the carcinogenic
potential of potassium chloride by
consulting the:
A. Chemical hygiene plan
B. Material safety data sheets
C. OSHA standards
D. Urinalysis procedure manual
14. Employees should not work with
radioisotopes if they are:
A. Wearing contact lenses
B. Allergic to iodine
C. Sensitive to latex
D. Pregnant
15. All of the following are safe to do when
removing the source of an electric shock
except:
A. Pulling the person away from the
instrument B. Turning off the circuit
breaker
C. Using a glass container to move the
instrument D. Unplugging the instrument
16. The acronym PASS refers to:
A. Presence of vital chemicals
B. Operation of a fire extinguisher
 C. Labeling of hazardous material B. Class B
D. Presence of radioactive substances C. Class C
17. The system used by firefighters when a D. Class D
fire occurs in the laboratory is: 24. Employers are required to provide free
A. MSDS immunizaton for:
B. RACE A. HIV
C. NFPA B. HTLV-1
D. PASS C. HBV
D. HCV
Continued on following page 25. A possible physical hazard in the
©2008 F. A. Davis hospital is: A. Wearing closed-toed
shoes
10 CHAPTER 1 • Safety in the Clinical Laboratory B. Not wearing jewelry
C. Having short hair
D. Running to answer the telephone
©2008 F. A. Davis
Continued
18. A class ABC fire extinguisher contains:
A. Sand
B. Water
C. Dry chemicals
D. Acid
19. The first thing to do when a fire is
discovered is to: A. Rescue persons in
danger
B. Activate the alarm system
C. Close doors to other areas
D. Extinguish the fire if possible
20. If a red rash is observed after removing
gloves, the employee:
Renal
Function
A. May be washing her hands too often
B. May have developed a latex allergy
C. Should apply cortisone cream
D. Should not rub the hands so vigorously
21. Pipetting by mouth is:
A. Acceptable for urine but not serum
B. Not acceptable without proper training
C. Acceptable for reagents but not specimens LEARNING OBJECTIVES
D. Not acceptable in the laboratory Upon completion of this chapter, the reader will be able
to:

22. The NPFA classification symbol contains

information on all of the following except:
A. Fire hazards
B. Biohazards
C. Reactivity
R

T
2
D. Health hazards P

23. The classification of a fire that can be H

extinguished with water is: C

A. Class A

1 Identify the components of the nephron,
kidney, and excretory system.
2 Trace the flow of blood through the nephron
and state the physiologic functions that occur. 3
Describe the process of glomerular ultrafiltration.
4 Discuss the functions and regulation of the
renin angiotensin-aldosterone system.
5 Differentiate between active and passive
transport in relation to renal concentration.
6 Explain the function of antidiuretic hormone
in the concentration of urine.
7 Describe the role of tubular secretion in
main taining acid-base balance.
8 Identify the laboratory procedures used to
evalu ate glomerular filtration, tubular
reabsorption and secretion, and renal blood
flow.
9 Discuss the advantages and disadvantages
active transport
aldosterone
maximal reabsorptive capacity
osmolarity
passive transport
podocytes
renal threshold
renal tubular acidosis
renin
renin-angiotensin-aldosterone
system 11
titratable acidity tubular
©2008 F. A. Davis
12 CHAPTER 2 • Renal Function
This chapter reviews nephron anatomy and
physiology and discusses their relationship to
urinalysis and renal function testing. A section on
laboratory assessment of renal function is included.
■■● Renal Physiology
Each kidney contains approximately 1 to 1.5 million
func tional units called nephrons. As shown in Figure
in using urea, inulin, creatinine, beta2
microglobu
lin, cystatin C, and
radionucleotides to measure glomerular
filtration.
KEY TERMS
10 Given hypothetic laboratory data, calculate a
cre atinine clearance and determine whether
the result is normal.
11 Discuss the clinical significance of the
creatinine clearance test.
12 Given hypothetic laboratory data, calculate
an estimated glomerular filtration rate.
13 Define osmolarity and discuss its
relationship to urine concentration.
14 Describe the basic principles of
clinical osmometers.
15 Given hypothetic laboratory data,
calculate a free-water clearance and
interpret the result.
16 Given hypothetic laboratory data,
calculate a PAH clearance and relate
this result to renal blood flow.
17 Describe the relationship of urinary
ammonia and titratable acidity to the
production of an acidic urine.
reabsorption tubular secretion
vasopressin
2-1, the human kidney contains two types of
nephrons. Cortical nephrons, which make up
approximately 85% of nephrons, are situated primarily
in the cortex of the kidney. They are responsible
primarily for removal of waste products and reab
sorption of nutrients. Juxtamedullary nephrons have
longer
Glomerulus
 The renal artery supplies blood to the kidney. The
human kid neys receive approximately 25% of the
blood pumped

loops of Henle that extend deep into the medulla of the

kid ney. Their primary function is concentration of the
urine. The ability of the kidneys to clear waste products
selec tively from the blood and simultaneously to
maintain the body’s essential water and electrolyte
balances is controlled in the nephron by the following
renal functions: renal blood flow, glomerular filtration, Renal
cortex
tubular reabsorption, and tubular secretion. The
physiology, laboratory testing, and associated
pathology of these four functions are discussed in this Cortical
chapter.

Renal Blood Flow

medulla

Loop of

Papilla of pyramid

Renal nephron tubule

Renal
Henle

Collecting
Renal duct
artery

Renal
vein
Calyx

Cortex

Renal pelvis
Juxtamedullary
nephron

Figure 2–1 The relationship of the

nephron to
the kidney and excretory system. (From
Scan
lon,VC, and Sanders,T: Essentials of
Ureter Anatomy
and Physiology, ed 3. FA Davis,
Philadelphia,

Urinary bladder
Urethra
1999, p 405, with permission.)
©2008 F. A. Davis

through the heart at all times. Blood enters the

capillaries of the nephron through the afferent
arteriole. It then flows through the glomerulus and
into the efferent arteriole. The varying sizes of these
arterioles help to create the hydrostatic pressure
differential important for glomerular filtration and to
maintain consistency of glomerular capillary pressure
and renal blood flow within the glomerulus. Notice the
smaller size of the efferent arteriole in Figure 2-2.
This increases the glomerular capillary pressure.
Before returning to the renal vein, blood from the
effer ent arteriole enters the peritubular capillaries
and the vasa recta and flows slowly through the
cortex and medulla of the kidney close to the tubules.
The peritubular capillaries surround the proximal and
distal convoluted tubules, provid ing for the
immediate reabsorption of essential substances from
the fluid in the proximal convoluted tubule and final
adjustment of the urinary composition in the distal
convo luted tubule. The vasa recta are located
adjacent to the ascending and descending loop of
Henle in juxtamedullary nephrons. In this area, the
major exchanges of water and salts take place
between the blood and the medullary inter stitium.
This exchange maintains the osmotic gradient (salt
concentration) in the medulla, which is necessary for
renal concentration.
Based on an average body size of 1.73 m2 of
surface, the total renal blood flow is approximately
1200 mL/min, and the total renal plasma flow
ranges 600 to 700 mL/min. Normal values for renal
blood flow and renal function tests depend on body
size. When dealing with sizes that vary greatly from
the average 1.73 m2 of body surface, a correction
must be calcu lated to determine whether the
observed measurements rep resent normal function.
This calculation is covered in the discussion on tests
for glomerular filtration rate (GFR) later
in this chapter. Variations in normal values have been
pub lished for different age groups and should be
considered when evaluating renal function studies.
Glomerular Filtration
The glomerulus consists of a coil of approximately
eight cap illary lobes referred to collectively as the
capillary tuft. It is located within Bowman’s capsule,
which forms the beginning of the renal tubule. Although
the glomerulus serves as a non selective filter of
plasma substances with molecular weights of less than
70,000, several factors influence the actual filtration
process. These include the cellular structure of the
capillary walls and Bowman’s capsule, hydrostatic and
oncotic pres sures, and the feedback mechanisms of
the renin angiotensin-aldosterone system. Figure
2-3 provides a diagrammatic view of the glomerular
areas influenced by these factors.
Cellular Structure of the Glomerulus
Plasma filtrate must pass through three cellular layers:
the capillary wall membrane, the basement membrane
(basal lamina), and the visceral epithelium of
Bowman’s capsule. The endothelial cells of the
capillary wall differ from those in other capillaries by
containing pores and are referred to as fenestrated.
The pores increase capillary permeability but do not
allow the passage of large molecules and blood cells.
Fur ther restriction of large molecules occurs as the
filtrate passes through the basement membrane and
the thin membranes covering the filtration slits formed
by the intertwining foot processes of the podocytes of
the inner layer of Bowman’s cap sule (see Fig. 2-3).

CHAPTER 2 • Renal Function 13

tubule

Collecting
Afferent
arteriole
Efferent
arteriole duct
Bowman’s capsule
Glomerulus Vasa recta
Cortex
Juxtaglomerular
Distal
Thick ascending
apparatus Proximal
convoluted
tubule

Peritubular
capillaries

Thick descending loop

of Henle Medulla
convoluted

Vasa recta loop of Henle

 component parts.
Thin descending loop of Henle
Thin ascending loop of Henle
Figure 2–2 The nephron and its
©2008 F. A. Davis

14 CHAPTER 2 • Renal Function

Afferent
arteriole

Efferent
arteriole

Hydrostatic
pressure

Oncotic
pressure
(unfiltered membrane
plasma
proteins) Bowman’s
space
Visceral
epithelium Proximal convoluted tubule
(podocyte) Figure 2–3 Factors affecting glomerular
Foot filtration in the renal corpuscle.
processes Endothelium
of podocyte

Basement
macula densa senses such changes, a cascade of
reactions within the RAAS occurs
Glomerular Pressure
As mentioned previously, the presence of hydrostatic
pressure resulting from the smaller size of the efferent
arteriole and the glomerular capillaries enhances
filtration. This pressure is necessary to overcome the
opposition of pressures from the fluid within Bowman’s
capsule and the oncotic pressure of unfiltered plasma
proteins in the glomerular capillaries. By increasing or
decreasing the size of the afferent arteriole, an
autoregulatory mechanism within the juxtaglomerular
appa ratus maintains the glomerular blood pressure at
a relatively constant rate regardless of fluctuations in
systemic blood pressure. Dilation of the afferent
arterioles and constriction of the efferent arterioles
when blood pressure drops prevent a marked decrease
in blood flowing through the kidney, thus preventing an
increase in the blood level of toxic waste prod ucts.
Likewise, an increase in blood pressure results in con
striction of the afferent arterioles to prevent
overfiltration or damage to the glomerulus.
Renin-Angiotensin-Aldosterone System
The renin-angiotensin-aldosterone system (RAAS)
controls the regulation of the flow of blood to and within
the glomeru lus. The system responds to changes in
blood pressure and plasma sodium content that are
monitored by the juxta glomerular apparatus, which
consists of the juxtaglomerular cells in the afferent
arteriole and the macula densa of the dis tal
convoluted tubule (Fig. 2-4). Low plasma sodium
content decreases water retention within the circulatory
system, resulting in a decreased overall blood volume
and subsequent decrease in blood pressure. When the
(Fig. 2-5). Renin, an enzyme produced by the
juxtaglomeru lar cells, is secreted and reacts with the
blood-borne substrate angiotensinogen to produce the
inert hormone angiotensin I. As angiotensin I passes
through the lungs, angiotensin con verting enzyme
(ACE) changes it to the active form angiotensin II.
Angiotensin II corrects renal blood flow in the following
ways: causing vasodilation of the afferent arte rioles
and constriction of the efferent arterioles, stimulating
reabsorption of sodium in the proximal convoluted
tubules, and triggering the release of the
sodium-retaining hormone aldosterone by the adrenal
cortex and antidiuretic hormone by the hypothalmus
(Table 2–1). As systemic blood pressure and plasma
sodium content increase, the secretion of renin
decreases. Therefore, the actions of angiotensin II
produce a constant pressure within the nephron.
As a result of the above glomerular mechanisms,
every minute approximately two to three million
glomeruli filter approximately 120 mL of
water-containing low-molecular weight substances.
Because this filtration is nonselective, the only
difference between the compositions of the filtrate and
the plasma is the absence of plasma protein, any
protein bound substances, and cells. Analysis of the
fluid as it leaves the glomerulus shows the filtrate to
have a specific gravity of 1.010 and confirms that it is
chemically an ultrafiltrate of plasma. This information
provides a useful baseline for eval uating the renal
mechanisms involved in converting the plasma
ultrafiltrate into the final urinary product.
Tubular Reabsorption
 the plasma ultrafil
The body cannot lose 120 mL of water-containing
essential substances every minute. Therefore, when
©2008 F. A. Davis

CHAPTER 2 • Renal Function 15

Distal tubule
Afferent
arteriole

Macula
Efferent
densa
arteriole

Juxtaglomerular
cells

Glomerulus

Figure 2–4 Close contact of the distal tubule with the affer
ent arteriole, macula densa, and the juxtaglomerular cells
within the juxtaglomerular apparatus.
Passive transport is the movement of molecules
across a membrane as a result of differences in their
trate enters the proximal convoluted tubule, the concentration or electrical potential on opposite sides
nephrons, through cellular transport mechanisms, of the membrane. These
begin reabsorbing these essential substances and
water (Table 2–2).
Low blood pressure
Low plasma sodium
Reabsorption Mechanisms
The cellular mechanisms involved in tubular
reabsorption are termed active and passive Renin secretion
transport. For active transport to occur, the substance
to be reabsorbed must combine with a carrier protein Angiotensinogen
contained in the membranes of the renal Angiotensin I
tubular cells. The electrochemical energy created by
this interaction transfers the substance across the cell Angiotensin
converting enzymes
mem branes and back into the bloodstream. Active
transport is responsible for the reabsorption of glucose, Angiotensin II
amino acids, and salts in the proximal convoluted
tubule, chloride in the ascending loop of Henle, and
sodium in the distal convo luted tubule. Proximal convoluted tubule
Vasoconstriction Aldosterone Sodium ADH
reabsorption

Collecting duct

Distal convoluted tubule

Figure 2–5 Algorithm of the renin
Water reabsorption angiotensin-aldosterone system. Sodium reabsorption

©2008 F. A. Davis

16 CHAPTER 2 • Renal Function

Table 2–1 Actions of the
RAAS 1. Dilation of the afferent arteriole and Reabsorption Substance Location
constriction of

Table 2–2 Tubular

the efferent arteriole the walls of which are
Active transport Passive
2. Stimulation of sodium impermeable to water. Urea
reabsorption in the proximal is passively reabsorbed in
convoluted tubule the proximal convo luted
3. Triggers the adrenal tubule and the ascending
cortex to release the loop of Henle, and passive Urea
sodium retaining hormone, reabsorption of sodium
aldosterone, to cause accompanies the active
reabsorp tion of sodium and transport of chloride in the transport
excretion of potassium in ascending loop.
Sodium
the distal convoluted tubule Active transport, like
and collecting duct passive transport, can be Proximal con voluted tubule
4. Triggers release of influ enced by the Ascending loop of Henle
antidiuretic hormone by the concentration of the Proximal and distal convo
hypo thalmus to stimulate substance being trans luted tubules
water reabsorption in the ported. When the plasma
col lecting duct concentration of a Glucose, amino acids, salts Proximal con voluted
substance that is normally Chloride tubule, descending
completely reabsorbed loop of Henle, and
physical differences are reaches an abnormally high collecting duct
Sodium
called gradients. Passive Proximal con voluted tubule
reabsorption of water takes and ascending loop of
place in all parts of the Henle
nephron except the Water Ascending loop of Henle
ascending loop of Henle,
Glucose, Na+
ple, glucose appearing in the urine of a person with a
level, the filtrate concentration exceeds the maximal normal blood glucose level is the result of tubular
reab sorptive capacity (Tm) of the tubules, and the damage and not diabetes mellitus.
substance begins appearing in the urine. The plasma Active transport of more than two-thirds of the
concentration at which active transport stops is termed filtered sodium out of the proximal convoluted tubule is
the renal threshold. For glucose, the renal threshold is accompa nied by the passive reabsorption of an equal
160 to 180 mg/dL, and glu cose appears in the urine amount of water. Therefore, as can be seen in Figure
when the plasma concentration reaches this level. 2-6, the fluid leaving the proximal convoluted tubule still
Knowledge of the renal threshold and the plasma maintains the same concen tration as the ultrafiltrate.
concentration can be used to distinguish between
excess solute filtration and renal tubular damage. For
exam

Aldosterone-controlled
Na+ reabsorption
Cortex
H2O

H2O

amino acids
H2O
ADH-controlled
H2O reabsorption

Na+

H2O
Cl–
 900 mOsm

300 mOsm

300 mOsm

600 mOsm Medulla

Figure 2–6 Renal
concentration.

1200 mOsm
©2008 F. A. Davis

Tubular Concentration
Renal concentration begins in the descending and
ascending loops of Henle, where the filtrate is
exposed to the high osmotic gradient of the renal
medulla. Water is removed by osmosis in the
descending loop of Henle, and sodium and chloride
are reabsorbed in the ascending loop. Excessive
reab sorption of water as the filtrate passes through
the highly con centrated medulla is prevented by the
water-impermeable walls of the ascending loop. This
selective reabsorption process is called the
countercurrent mechanism and serves to maintain
the osmotic gradient of the medulla. The sodium and
chloride leaving the filtrate in the ascending loop
prevent dilution of the medullary interstitium by the
water reabsorbed from the descending loop.
Maintenance of this osmotic gradi ent is essential for
the final concentration of the filtrate when it reaches
the collecting duct.
In Figure 2-6, the actual concentration of the
filtrate leaving the ascending loop is quite low owing
to the reab sorption of salt and not water in that part
of the tubule. Reab sorption of sodium continues in
the distal convoluted tubule, but it is now under the
control of the hormone aldosterone, which regulates
reabsorption in response to the body’s need for
sodium (see Fig. 2-5).

Collecting Duct Concentration

The final concentration of the filtrate through the
reabsorp tion of water begins in the late distal
convoluted tubule and continues in the collecting
duct. Reabsorption depends on the osmotic gradient
in the medulla and the hormone vasopressin
(antidiuretic hormone [ADH]). One would expect that
as the dilute filtrate in the collecting duct comes in
contact with the higher osmotic concentration of the
 medullary interstitium, passive reabsorption of water
would occur. However, the process is controlled by the
presence or absence of ADH, which renders the walls
of the distal convoluted tubule and collecting duct
permeable or imper meable to water. A high level of
ADH increases permeability, resulting in increased
reabsorption of water, and a low volume concentrated
urine. Likewise, absence of ADH ren ders the walls
impermeable to water, resulting in a large volume of
dilute urine. Just as the production of aldosterone is
controlled by the body’s sodium concentration, produc
tion of ADH is determined by the state of body
hydration. Therefore, the chemical balance in the body
is actually the final determinant of urine volume and
concentration. The concept of ADH control can be
summarized in the following manner:
↑Body Hydration ↓ADH ↑Urine Volume
↓Body Hydration ↑ADH ↓Urine Volume

Tubular Secretion
In contrast to tubular reabsorption, in which substances
are removed from the glomerular filtrate and returned
to the blood, tubular secretion involves the passage of
substances from the blood in the peritubular capillaries
to the tubular fil trate (Fig. 2-7). Tubular secretion
serves two major functions: elimination of waste
products not filtered by the glomerulus and regulation
of the acid-base balance in the body through the
secretion of hydrogen ions.
Many foreign substances, such as medications,
cannot be filtered by the glomerulus because they are
bound to plasma proteins. However, when these
protein-bound substances enter the peritubular
capillaries, they develop a stronger affin

Peritubular
capillary

Bowman’s
capsule
To blood Reabsorption
CHAPTER 2 • Renal Function 17
substances in the nephron.
Glomerular To urine
filtrate Secretion

Figure 2–7 The movement of Tubule

©2008 F. A. Davis

18 CHAPTER 2 • Renal Function

 lumen Renal tubular cell lumen HPO4 (filtered) Peritubular plasma
Tubular Peritubular plasma – Renal tubular cell
–
HCO3 (filtered) Tubular
H+ + H+ HCO3– HCO3– HPO HCO3– HCO3– H2CO3
4
+ –+
H2CO3 H +H
HCO3
Carbonic anhydrase H2O +CO2 H2PO4
H2CO3 Carbonic anhydrase

H2O + CO2

H2O + CO2 CO2 Final urine Final urine

CO2
diagrams of the two
Figure 2–9 Excretion of secreted hydrogen ions
Figure 2–8 Reabsorption of filtered bicarbonate. combined with phosphate.

■■● Renal Function Tests

ity for the tubular cells and dissociate from their carrier This brief review of renal physiology shows that there
pro teins, which results in their transport into the filtrate are many metabolic functions and chemical interactions
by the tubular cells. The major site for removal of these to be evaluated through laboratory tests of renal
nonfiltered substances is the proximal convoluted function. In Figure 2-11, the parts of the nephron are
tubule. related to the laboratory tests used to assess their
function.
Acid-Base Balance
To maintain the normal blood pH of 7.4, the blood must Glomerular Filtration Tests
buffer and eliminate the excess acid formed by dietary
intake and body metabolism. The buffering capacity of The standard test used to measure the filtering
- capacity of the glomeruli is the clearance test. As its
the blood depends on bicarbonate (HCO3 ) ions, which
name implies, a clearance test measures the rate at
are readily filtered by the glomerulus and must be
which the kidneys are able to remove (to clear) a
expedi ently returned to the blood to maintain the
filterable substance from the blood. To ensure that
proper pH. As shown in Figure 2-8, the secretion of
glomerular filtration is being measured accurately, the
hydrogen ions (H ) by the renal tubular cells into the
substance analyzed must be one that is neither
filtrate prevents the filtered bicarbonate from being reabsorbed nor secreted by the tubules. Other factors
excreted in the urine and causes the return of a to consider in the selection of a clearance test
bicarbonate ion to the plasma. This process provides substance include the stability of the substance in urine
for almost 100% reabsorption of filtered bicarbonate during a possible 24-hour collection period, the
and occurs primarily in the proximal convoluted tubule. consistency of the plasma level, the substance’s
As a result of their small molecular size, hydrogen availability to the body, and the availability of tests for
ions are readily filtered and reabsorbed. Therefore, the analy sis of the substance.
actual excretion of excess hydrogen ions also depends
on tubular secretion. Figures 2-9 and 2-10 are
tubule, ammonia is pro resulting ammonium ion is
NH3
Peritubular plasma excreted in the urine.
primary methods for NH 3 All three of these
+ +
hydrogen ion excretion in H + H processes occur
the urine. In Figure 2-9 the rather than reabsorbed. simultaneously at
Additional excretion of HCO3– HCO3– H2CO3
secreted hydrogen ion
hydrogen ions is NH4
combines with a filtered duced from the breakdown +
phosphate ion instead of a accomplished through of the amino acid Carbonic anhydrase
bicarbonate ion and is their reaction with
glutamine. The ammonia
excreted ammonia pro duced and
reacts with the H to form
Tubular lumen secreted by the cells of the
Renal tubular cell distal convoluted tubule. In the ammonium ion (NH4 ) H2O + CO2
the proximal convoluted (see Fig. 2-10). The CO2

Final urine
rates determined by the acid-base balance in the body.
A dis ruption in these secretory functions can result in Figure 2–10 Excretion of secreted hydrogen ions
metabolic acidosis or renal tubular acidosis, the combined with ammonia produced by the tubules.
inability to produce an acid urine.
©2008 F. A. Davis
 acidity
PAH

Clearance
tests Free water
clearance

CHAPTER 2 • Renal Function 19 Osmolarity

Ammonia
Titratable

Figure 2–11 The relationship of nephron areas
to renal function tests.
Clearance Tests
The earliest glomerular filtration tests measured urea
because of its presence in all urine specimens and the
existence of rou tinely used methods of chemical
analysis. Because approxi mately 40% of the filtered
urea is reabsorbed, normal values were adjusted to
reflect the reabsorption, and patients were hydrated to
produce a urine flow of 2 mL/min to ensure that no
more than 40% of the urea was reabsorbed. At
present, the use of urea as a test substance for
glomerular filtration has been replaced by the
measurement of other substances including
creatinine, inulin, beta2 microglobulin, cystatin C, or
radioisotopes. Each procedure has its advantages and
dis advantages.
Inulin Clearance
Inulin, a polymer of fructose, is an extremely stable
substance that is not reabsorbed or secreted by the
tubules. It is not a normal body constituent, however,
and must be infused at a constant rate throughout the
testing period. A test that requires an infused
substance is termed an exogenous proce dure and is
seldom the method of choice if a suitable test sub
stance is already present in the body (endogenous
procedure). Therefore, although inulin was the original
reference method for clearance tests, it is currently not
used for glomerular fil tration testing.
Creatinine Clearance
Currently, routine laboratory measurements of GFR
employ creatinine as the test substance. Creatinine, a
waste product of muscle metabolism that is normally
found at a relatively con stant level in the blood,
provides the laboratory with an endogenous procedure
for evaluating glomerular function. The use of creatinine ©2008 F. A. Davis
has several disadvantages not found with inulin, and
careful consideration should be given to them. They are
as follows:
Osmolarity
1. Some creatinine is secreted by the
tubules, and secretion increases as blood
levels rise.
2. Chromogens present in human plasma react
in the chemical analysis. Their presence,
however, may help counteract the falsely
elevated rates caused by tubular secretion.
3. Medications, including gentamicin,
cephalosporins, and cimetidine (Tagamet),
inhibit tubular secretion of creatinine, thus
causing falsely low serum levels.1
4. Bacteria will break down urinary creatinine if
speci mens are kept at room temperature for
extended periods.2
5. A diet heavy in meat consumed during
collection of a 24-hour urine specimen will
influence the results if the plasma specimen is
drawn prior to the collection period.
6. Measurement of creatinine clearance is not a
reliable indicator in patients suffering from
muscle-wasting diseases.
Because of these drawbacks, abnormal results
may be followed by more sophisticated tests, but the
creatinine clear ance test provides the clinical
laboratory with a method for screening the GFR.
Procedure
By far the greatest source of error in any clearance
procedure utilizing urine is the use of improperly timed
urine speci mens. The importance of using an
accurately timed specimen (see Chapter 3) will become
evident in the following discus sion of the calculations
involved in converting isolated labo ratory
measurements to GFR. The GFR is reported in
mL/min; therefore, determining the number of milliliters
of plasma from which the clearance substance
(creatinine) is completely removed during 1 minute is
necessary. To calculate this infor
20 CHAPTER 2 • Renal Function
mation, one must know urine volume in mL/min (V), urine
creatinine concentration in mg/dL (U), and plasma
creatinine concentration in mg/dL (P).
The urine volume is calculated by dividing the number of
milliliters in the specimen by the number of minutes used
to collect the specimen.
account variations in size and muscle mass. Values are
formula is:
Example
Calculate the urine volume (V) for a 2-hour
speci men measuring 240 mL:
2 hours 60 minutes 120 minutes
240 mL/120 minutes 2 mL/min
V 2 mL/min
The plasma and urine concentrations are
determined by chemical testing. The standard
formula used to calculate the milliliters of
plasma cleared per minute (C) is:
U
C PV
This formula is derived as follows. The
milliliters of plasma cleared per minute (C)
times the mg/dL of plasma creatinine (P) must
equal the mg/dL of urine creatinine (U) times
the urine volume in mL/min (V), because all of
the filtered creatinine will appear in the urine.
Therefore:
U Plasma filtrate (120 mL/min 0.01 mg/mL = 1.2 mg)
CP UV and C PV
Example
Using urine creatinine of 120 mg/dL (U),
plasma creatinine of 1.0 mg/dL (P), and
urine volume of 1440 mL obtained from a
24-hour specimen (V), calculate the GFR.
1440 mL
V 1 mL/min 60 minutes 24 1440 minutes
120 mg/dL (U) 1 mL/min (V)
C 120 mL/min 1.0 mg/dL (P)
By analyzing this calculation and referring to
Figure 2-12, at a 1 mg/dL concentration, each
milliliter of plasma contains 0.01 mg creatinine.
Therefore, to arrive at a urine concentration of
120 mg/dL (1.2 mg/mL), it is necessary to clear
120 mL of plasma. Although the filtrate volume
is reduced, the amount of creatinine in the
filtrate does not change because the creatinine
is not reabsorbed. Knowing that in the average
person (1.73 m2 body surface) the approximate
amount of plasma filtrate produced per minute
is 120 mL, it is not surprising that normal
creatinine clearance values approach 120
mL/min (men, 107 to 139 mL/min; women, 87
to 107 mL/min). The normal plasma creatinine
is 0.5 to 1.5 mg/dL. These normal values take
into
considerably lower in older people, however, and an
adjustment may also have to be made to the calculation
when dealing with body sizes that devi ate greatly from
1.73 m2 of surface, such as with children. To adjust a
clearance for body size, the
U 1. 73
C PV A
with A being the actual body size in square
meters of surface. The actual body size may be
calculated as:
log A (0.425 log weight)
(0.725 log height) -2.144
or it may be obtained from the nomogram
shown in Figure 2-13.
Plasma (1 mg/dL = 0.01 mg/mL creatinine)
Glomerulus
Reabsorption
(119 mL H 2O)
Urine (1 mL/min)
Creatinine (1.2 mg/mL or 120 mg/dL)
Figure 2–12 A diagram representing creatinine filtration
and excretion.
Clinical Significance
When interpreting the results of a creatinine clearance
test, the GFR is determined not only by the number of
functioning nephrons but also by the functional
capacity of these nephrons. In other words, even
though half of the available nephrons may be
nonfunctional, a change in the GFR will not occur if the
remaining nephrons double their filtering capacity. This
is evidenced by persons who lead normal lives with
only one kidney. Therefore, although the creatinine
clearance is a frequently requested laboratory
procedure, its value does not lie in the detection of
early renal disease. Instead, it is used to determine the
extent of nephron damage in known cases of renal
disease, to monitor the effectiveness of treatment
designed to prevent further nephron damage, and to
determine the feasibility of administering medications,
which can build up to dangerous blood levels if the
GFR is markedly reduced.
Calculated Glomerular Filtration Estimates
Formulas have been developed to provide estimates of
 the GFR based on the serum creatinine without the urine creati
©2008 F. A. Davis 400
180

Ccr
CHAPTER 2 • Renal Function 21

200
440 (140 - age)(weight in kilograms)
420
190 72 serum creatinine in mg/dL
dn
u may not be ) gi

e albumin. black) infused

.60
BUN-0.170 substances
o

380 360 340 320 the result Several

H

2"
300 290 280 270 of muscle A newer .55 vari serum , thereby
4'
mass, i.e., formula, 10" 8"
i
260 250 240 230 k
albumin eliminating
220 210 200 190 .50
180 170 160 150 fatty tissue. called the 0.318 the need
n
p
i

Modificatio 6"
n

140 130 120 The i

t
for urine
170 160 150 140 calculation n of Diet in 4"
h

A collection,
ations of
gi

130 120 110 for ideal Renal 2" laboratory has

W

110 100 the formula

body Disease 3' advantage enhanced
100 95 are
weight (MDRD) of this inter est in
90
available,
85 (IBW) is: system, 10" 8" formula is exogenous
utilizing
80 Males: 50 utilizes one or that, as procedures
75 kg 2.3 kg additional 6" body . Injection
125 120 115 110
more of
s
70 for each variables 105 100 95
s
90 t

h
weight is of
r

e
65 inch of
m
g
i
omitted, all radionucle
80
height over 90
results are otides such
te

m
te 60
e
e 60 inches 85 available as
55 70
r

Females: from the 125I-iothala

f

7'
a
the
The results 45.5 kg
u

10" 8" 80
6"
q

3.00 2.90 2.80 additional labora tory mate

4" 2.70 2.60 2.50 are 2.3 kg for 60
variables. computer provides a
75
2" 2.40 2.30 2.20 multiplied each inch n

An information method for

2.10
e
6'
10" 8" 2.00 1.95 1.90
by 0.85 for of height c

example of , and the determinin

6" 1.85 1.80 1.75 female over 60
n
i
50
the MDRD calculation g
1.70 1.65 1.60
4"
1.55 1.50 1.45 patients. inches t

h study for can be per glomerular

2"
1.40 1.35 1.30 Modificatio The gi

e
e

formed filtration
5' 1.25 W

10" 8"
ns to the H
40
automatica through the
s
50 45 40
6"
original calculation n
i
lly by the plasma
4" formula a

e instrument disappeara
substitute for
220 215 210 205 r 35 30
200 195 190 185
a
mula is: performing nce of the
180 175 170 165
ideal body e
the serum
weight in adjusted
c

160 155 150 145 a

GFR 170 creatinine.4
25 20
and does
fr

140 135 130 kilograms u

serum The
sr
body not include 1.20 1.15 1.10
S

e and creatinine-0 developme

body
te
1.05 1.00 .95
m
it
adjusted weight .999
nt of
body weight. .90
age-0.176 simplified
weight in (AjBW) is: The .85

variables .80
15
0.822 (if procedures
kilo grams.
l

nicity, patient is measuring

This is IBW 0.3 include eth .75
a
blood urea female) the plasma
done to r
.70
g
nitrogen, 1.1880 ( if disappeara
correct for (ABW-IBWo

n
.65 and serum patient is nce of
weight that
i
t

s h

Figure 2–13 A nomogram for the determination of body
surface area. (From Boothby,WM, and Sandiford, RB:
Nomo gram for determination of body surface area. N
Engl J Med 185:227, 1921, with permission.)
nine. These formulas are becoming more frequently
used in clinical medicine. As discussed, the creatinine
clearance is not useful in detecting early renal disease.
Therefore the calcu lated clearances are being used for
monitoring patients already diagnosed with renal
disease or at risk for renal dis ease. In addition, the
formulas are valuable when medications that require
adequate renal clearance need to be prescribed.
The most frequently used formula was developed
by Cockcroft and Gault.3 It is also used to document
eligibility for reimbursement by the Medicare End Stage
Renal Disease Program and for evaluating patient
placement on kidney transplant lists.4 Variables
included in the original formula are age, sex, and body
weight in kilograms.
radioactive material and enables visualization of the
filtration in one or both kidneys.5
Good correlation between the GFR and plasma
levels of beta2 microglobulin has been demonstrated.
microglob
Beta2 ulin (molecular weight 11,800)
dissociates from human leukocyte antigens at a
constant rate and is rapidly removed from the plasma
by glomerular filtration. Sensitive methods using
 enzyme immunoassay are available for the measure
ment of beta2 microglobulin.6 A rise in the plasma level
of beta2 microglobulin has been shown to be a more
sensitive indicator of a decrease in GFR than creatinine
clearance. However, the test is not reliable in patients
who have a history of immunologic disorders or
malignancy.7
Another serum marker that can be used to monitor
GFR is cystatin C. Cystatin C is a small protein
(molecular weight 13,359) produced at a constant rate
by all nucleated cells. It is readily filtered by the
glomerulus and reabsorbed and broken down by the
renal tubular cells. Therefore, no cystatin C is secreted
by the tubules, and the serum concentration can
©2008 F. A. Davis
22 CHAPTER 2 • Renal Function
be directly related to the GFR. Immunoassay
procedures are available for measuring cystatin
C.8 Monitoring levels of cys tatin C is
recommended for pediatric patients, persons with
diabetes, the elderly, and critically ill patients.9
Tubular Reabsorption Tests
Whereas measurement of the GFR is not a useful
indication of early renal disease, the loss of tubular
reabsorption capability is often the first function
affected in renal disease. This is not surprising
when one considers the complexity of the tubular
reabsorption process.
Tests to determine the ability of the tubules to
reabsorb the essential salts and water that have
been nonselectively filtered by the glomerulus are
called concentration tests. As mentioned, the
ultrafiltrate that enters the tubules has a specific
gravity of 1.010; therefore, after reabsorption one
would expect the final urine product to be more
concen trated. However, as you perform routine
urinalysis, you will see that many specimens do
not have a specific gravity higher than 1.010, yet
no renal disease is present. This is because urine
concentration is largely determined by the body’s
state of hydration, and the normal kidney will
reabsorb only the amount of water necessary to
preserve an adequate supply of body water.
As can be seen in Figure 2-14, both
specimens contain the same amount of solute;
however, the urine density (spe cific gravity) of
patient A will be higher. Therefore, control of fluid
intake must be incorporated into laboratory tests
that measure the concentrating ability of the
kidney.
Throughout the years, various methods have
been used to produce water deprivation, including
the Fishberg and Mosenthal concentration tests,
Patient A
Glomerulus
Water (1 glass)
which measured specific gravity. In the Fishberg
test, patients were deprived of fluids for 24 hours
prior to measuring specific gravity. The Mosen thal
test compared the volume and specific gravity of
day and night urine samples to evaluate
concentrating ability. Neither test is used now
because the information provided by specific
gravity measurements is most useful as a screening
procedure, and quantitative measurement of renal
concentrating ability is best assessed through
osmometry. However, persons with normal
concentrating ability should have a specific gravity
of 1.025 when deprived of fluids for 16 hours.
Following overnight water deprivation, a urine
osmolarity of 800 mOsm or above indicates normal
concentrating ability.
Osmolarity
Specific gravity depends on the number of particles
present in a solution and the density of these
particles, whereas osmo larity is affected only by the
number of particles present. When evaluating renal
concentration ability, the substances of interest are
small molecules, primarily sodium (molecular
weight, 23) and chloride (molecular weight, 35.5).
However, urea (molecular weight, 60), which is of
no importance to this evaluation, will contribute
more to the specific gravity than will the sodium and
chloride molecules. Because all three molecules
contribute equally to the osmolarity of the speci
men, a more representative measure of renal
concentrating ability can be obtained by measuring
osmolarity.
An osmole is defined as 1 g molecular weight
of a sub stance divided by the number of particles
into which it disso ciates. A nonionizing substance
such as glucose (molecular weight, 180) contains
180 g per osmole, whereas sodium chloride (NaCl)
(molecular weight, 58.5), if completely dis sociated,
contains 29.25 g per osmole. Just like molality and
molarity, there are osmolality and osmolarity. An
osmolal solution of glucose has 180 g of glucose
dissolved in 1 kg of solvent. An osmolar solution of
glucose has 180 g of glucose dissolved in 1 L of
solvent. In the clinical laboratory, the terms are used
interchangeably, inasmuch as the difference in nor
mal temperature conditions with water as the
solvent is min imal. The unit of measure used in the
clinical laboratory is the milliosmole (mOsm),
because it is not practical when dealing with body
fluids to use a measurement as large as the osmole
(23 g of sodium per liter or kilogram).
Ultrafiltrate 120 mL Water
300 mg Solute
Reabsorption 119 mL Water
100 mg Solute
 1 mL Water
200 mg Solute Urine 200 mg Solute Urine
Specific gravity 1.015 Specific gravity 1.005
Patient B
Water (4 glasses)
Glomerulus
Ultrafiltrate 120 mL Water
300 mg Solute
Reabsorption 110 mL Water
100 mg Solute
©2008 F. A. Davis
The osmolarity of a solution can be determined
by measuring a property that is mathematically
related to the number of particles in the solution
(colligative property) and comparing this value with
the value obtained from the pure solvent. Solute
dissolved in solvent causes the following changes in
colligative properties: lower freezing point, higher
boiling point, increased osmotic pressure, and lower
vapor pressure.
Because water is the solvent in both urine and
plasma, the number of particles present in a sample
can be deter mined by comparing a colligative
property value of the sam ple with that of pure water.
Clinical laboratory instruments are available to
measure freezing point depression and vapor
pressure depression.
Freezing Point Osmometers
Measurement of freezing point depression was the
first prin ciple incorporated into clinical osmometers,
and many instru ments employing this technique are
available. These osmometers determine the freezing
point of a solution by supercooling a measured
amount of sample to approximately 27 C. The
supercooled sample is vibrated to produce crystal
lization of water in the solution. The heat of fusion
produced by the crystallizing water temporarily raises
the temperature of the solution to its freezing point. A
temperature-sensitive probe measures this
temperature increase, which corresponds to the
freezing point of the solution, and the information is
converted into milliosmoles. Conversion is made
possible by the fact that 1 mol (1000 mOsm) of a
nonionizing substance dissolved in 1 kg of water is
known to lower the freezing point 1.86 C. Therefore,
by comparing the freezing point depression of an
unknown solution with that of a known molal solution,
the osmolarity of the unknown solution can be
calculated. Clinical osmometers use solutions of
known NaCl concentration as their reference
standards because a solution of partially ionized
10 mL Water
Figure 2–14 The effect of
hydration on renal concentration.
substances is more representative of urine and
plasma composition.
Vapor Pressure Osmometers
The other instrument used in clinical osmometry is
called the vapor pressure osmometer. The actual
measurement per formed, however, is that of the dew
point (temperature at which water vapor condenses
to a liquid). The depression of dew point temperature
by solute parallels the decrease in vapor pressure,
thereby providing a measure of this colligative
property.
Samples are absorbed into small filter paper
disks that are placed in a sealed chamber containing
a temperature sensitive thermocoupler. The sample
evaporates in the cham ber, forming a vapor. When
the temperature in the chamber is lowered, water
condenses in the chamber and on the ther
mocoupler. The heat of condensation produced
raises the temperature of the thermocoupler to the
dew point tempera ture. This dew point temperature
is proportional to the vapor pressure from the
evaporating sample. Temperatures are com pared
with those of the NaCl standards and converted into
CHAPTER 2 • Renal Function 23
milliosmoles. The vapor pressure osmometer uses
microsam ples of less than 0.01 mL; therefore, care
must be taken to prevent any evaporation of the
sample prior to testing. Corre lation studies have
shown more variation with vapor pressure
osmometers, stressing the necessity of careful
technique.
Technical Factors
Factors to consider because of their influence on true
osmolarity readings include lipemic serum, lactic acid,
and volatile substances, such as ethanol, in the
specimen. In lipemic serum, the displacement of serum
water by insoluble lipids produces erroneous results
with both vapor pres sure and freezing point
osmometers. Falsely elevated values owing to the
formation of lactic acid also occur with both methods if
serum samples are not separated or refrigerated within
20 minutes. Vapor pressure osmometers do not detect
the presence of volatile substances, such as alcohol,
as they become part of the solvent phase; however,
measurements per formed on similar specimens using
freezing point osmometers will be elevated.
Clinical Significance
Major clinical uses of osmolarity include initially
evaluating renal concentrating ability, monitoring the
course of renal disease, monitoring fluid and electrolyte
therapy, establishing the differential diagnosis of
hypernatremia and hypona tremia, and evaluating
the secretion of and renal response to ADH. These
evaluations may require determination of serum in
addition to urine osmolarity.
Normal serum osmolarity values are from 275 to
300 mOsm. Normal values for urine osmolarity are
difficult to establish, because factors such as fluid
intake and exercise can greatly influence the urine
concentration. Values can range from 50 to 1400
mOsm.2 Determining the ratio of urine to serum
osmolarity can provide a more accurate evaluation.
Under normal random conditions, the ratio of urine to
serum osmolarity should be at least 1:1; after
controlled fluid intake, it should reach 3:1.
The ratio of urine to serum osmolarity, in
conjunction with procedures such as controlled fluid
intake and injection of ADH, is used to differentiate
whether diabetes insipidus is caused by decreased
ADH production or inability of the renal tubules to
Uos
Cosm m
P os
and then subtracting the osmolar clearance value from
the urine volume in mL/min.
Example
Using a urine osmolarity of 600 mOsm (U), a
urine volume of 2 mL/min (V), and a plasma
osmolarity of 300 mOsm (P), calculate the free
water clearance: Cosm
600
3 0
0 2 (V)
4.0 mL/min peritubular
CH2O 2 (V) - 4.0 (Cosm) -2.0 (free water
clearance)
Calculation of the osmolar clearance indicates how
much water must be cleared each minute to produce a
urine with the same osmolarity as the plasma. The
ultrafiltrate con tains the same osmolarity as the
plasma; therefore, the osmotic differences in the urine
are the result of renal con centrating and diluting
mechanisms. By comparing the osmo lar clearance
respond to ADH. Failure to achieve a ratio of 3:1
following injection of ADH indicates that the collecting
duct does not have functional ADH receptors. In
contrast, if concentration takes place following ADH
injection, an inabil ity to produce adequate ADH is
indicated. Tests to measure the ADH concentration in
plasma and urine directly are avail able for difficult
diagnostic cases.10
Free Water Clearance
The ratio of urine to serum osmolarity can be further
expanded by performing the analyses using water
deprivation and a timed urine specimen and calculating
the free water clearance. The free water clearance is
determined by first
©2008 F. A. Davis
24 CHAPTER 2 • Renal Function
calculating the osmolar clearance using the standard
clear ance formula:
however, because of interference by medications and
elevated waste products in patients’ serum and the
necessity to obtain
V urine specimens. Therefore,
the
several very accurately timed
PSP test is not currently performed.
m
PAH Test
To measure the exact amount of blood flowing through
the kidney, it is necessary to use a substance that is
completely removed from the blood (plasma) each time
it comes in con tact with functional renal tissue. The
principle is the same as in the clearance test for
glomerular filtration. However, to ensure measurement
of the blood flow through the entire nephron, the
substance must be removed from the blood pri
(U
) (P)
marily in the capillaries rather blood reaches the
than being glomerulus.
removed when the
with the actual urine volume excreted per
Although it has the disadvantage of being
exogenous, the chemical PAH meets the criteria
needed to measure renal blood flow. This nontoxic
substance is loosely bound to plasma pro teins, which
permits its complete removal as the blood passes
through the peritubular capillaries. Except for a small
amount of PAH contained in plasma that does not
come in contact with functional renal tissue, all the
plasma PAH is secreted by the proximal convoluted
tubule. Therefore, the volume of plasma flowing
 through the kidneys determines the amount of PAH
excreted in the urine. The standard clearance formula
V (mL/min urine)
minute, it can be determined needed to maintain
whether the water being excreted
is more or less than the amount CPAH (mL/min)
an osmolarity the same as that of the ultrafiltrate. The
above calculation shows a free water clearance of -2.0,
indicating that less than the necessary amount of water
is being excreted, a possible state of dehydration. If the
value had been 0, no renal concentration or dilution
would be tak ing place; likewise, if the value had been
©2008 F. A. Davis
2.0, excess water would have been excreted.
Therefore, calculation of the free water clearance is
used to determine the ability of the kidney to respond to
the state of body hydration.
Tubular Secretion and
Renal Blood Flow Tests
Tests to measure tubular secretion of nonfiltered
substances and renal blood flow are closely related in
that total renal blood flow through the nephron must be
measured by a substance that is secreted rather than
filtered through the glomerulus. Impaired tubular
secretory ability or inadequate presentation of the
substance to the capillaries owing to decreased renal
blood flow may cause an abnormal result. Therefore,
an understanding of the principles and limitations of the
tests and correlation with other clinical data is impor
tant in test interpretation.
The test most commonly associated with tubular
secre tion and renal blood flow is the p-aminohippuric
acid (PAH) test. Historically, excretion of the dye
phenolsulfon phthalein (PSP) was used to evaluate
these functions. Stan dardization and interpretation of
PSP results are difficult,
can be used to calculate the effective renal plasma
flow. Based on normal hematocrit readings, normal
values for the effective renal plasma flow range from
600 to 700 mL/min, making the average renal blood
flow about 1200 mL/min. The actual measurement is
renal plasma flow rather than renal blood flow, because
the PAH is contained only in the plasma portion of the
blood. Also, the term “effective” is included because
approximately 8% of the renal blood flow does not
come into contact with the functional renal tissue.11
The amount of PAH infused must be monitored
carefully to ensure accurate results; therefore, the test
is usually per formed by specialized renal laboratories.
Nuclear medicine procedures using radioactive
hippurate can determine renal blood flow by measuring
the plasma disappearance of a sin gle radioactive
injection and at the same time provide visual ization of
the blood flowing through the kidneys.5
Titratable Acidity and Urinary Ammonia
The ability of the kidney to produce an acid urine
depends on the tubular secretion of hydrogen ions and
production and secretion of ammonia by the cells of the
distal convoluted tubule. A normal person excretes
approximately 70 mEq/day of acid in the form of either
U (mg/dL PAH)
P (mg/dL PAH)
titratable acid (H ), hydrogen phosphate ions (H2PO4-),
or ammonium ions (NH4 ). In normal persons, a diurnal
variation in urine acidity consisting of alkaline tides
appears shortly after arising and postprandi ally at
approximately 2 p.m. and 8 p.m. The lowest pH is
found at night.
The inability to produce an acid urine in the
presence of metabolic acidosis is called renal
tubular acidosis. This con dition may result from
impaired tubular secretion of hydro gen ions
associated with the proximal convoluted tubule or
defects in ammonia secretion associated with the
distal con voluted tubule.
Measurement of urine pH, titratable acidity,
and urinary ammonia can be used to determine
the defective function. The tests can be run
simultaneously on either fresh or toluene-pre
served urine specimens collected at 2-hour
intervals from patients who have been primed
with an acid load consisting of oral ammonium
chloride. By titrating the amount of free H
(titratable acidity) and then the total acidity of the
speci men, the ammonium concentration can be
calculated as the difference between the
titratable acidity and the total acidity.
References
1. Berger, A: Renal function and how to assess it.
Brit J Med 321:1444, 2000.
2. Pincus, MR, Preuss, HG, and Henry, JB:
Evaluation of renal function and water, electrolyte
and acid-base balance. In Henry, JB (ed): Clinical
Diagnosis and Management by Laboratory
Methods. WB Saunders, Philadelphia, 1996.
3. Cockcroft, DW, and Gault, HH: Prediction of
creatinine clear ance from serum creatinine. N Engl
J Med 281:1405-1415, 1969.
4. Levey, AS, et al: A more accurate method to
estimate glomeru lar filtration rate from serum
creatinine: A new prediction equa tion. Ann Intern
Med 130(6):461-470, 1999.
5. Chachati, A, et al: Rapid method for the
measurement of differential renal function:
Validation. J Nucl Med 28(5):
829-836, 1987.
6. Peterson, L: Beta2 microglobulin. Clin Chem
News 14(1):6, 1988.
7. Murray, B, and Ferris, TF: Blood and urinary
chemistries in the evaluation of renal function. Sem
Nephrol 5(3):208-221, 1985. 8. Laterza, OE, Price,
CP, and Scott, MG: Cystatin C: An improved
estimator of glomerular filtration rate? Clin Chem
48(5):699- 707, 2002.
9. Tan, GS, et al: Clinical usefulness of cystatin C for
the estima tion of glomerular filtration rate in type 1
diabetes. Crit Care 9(2):139-143, 2005.
 10. Daves, BB, and Zenser, TV: Evaluation of renal
concentrating and diluting ability. Clin Lab Med 8. The hormone aldosterone is
13(1):131-134, 1993. responsible for: A. Hydrogen ion
11. Duston, H, and Corcoran, A: Functional secretion
interpretation of renal tests. Med Clin North Am B. Potassium secretion
39:947-956, 1955.
C. Chloride retention
D. Sodium retention
STUDY
QUESTIONS 9. The fluid leaving the glomerulus has
a specific gravity of:
A. 1.005
B. 1.010
1. The type of nephron responsible for renal C. 1.015
concentra tion is the: D. 1.020
A. Cortical
B. Juxtaglomerular 10. All of the following are reabsorbed by active
2. The function of the peritubular capillaries is: transport in the tubules except:
A. Reabsorption A. Urea
B. Filtration B. Glucose
C. Secretion C. Sodium
D. Both A and C D. Chloride

Continued on following page

©2008 F. A. Davis
CHAPTER 2 • Renal Function 25

26 CHAPTER 2 • Renal Function

3. Blood flows through the nephron in the
following order:
A. Efferent arteriole, peritubular Continued
capillaries, vasa recta, afferent
arteriole 11. Which of the tubules is impermeable to
B. Peritubular capillaries, afferent arteriole, water? A. Proximal convoluted tubule
vasa recta, efferent arteriole B. Descending loop of Henle
C. Afferent arteriole, peritubular C. Ascending loop of Henle
capillaries, vasa recta, efferent D. Distal convoluted tubule
arteriole 12. Glucose will appear in the urine when the:
D. Efferent arteriole, vasa recta, peritubular A. Blood level of glucose is 200 mg/dL
capillar ies, afferent arteriole B. Tm for glucose is reached
C. Renal threshold for glucose is exceeded
4. Filtration of protein is prevented in the D. All of the above
glomerulus by:
A. Hydrostatic pressure 13. The countercurrent mechanism takes
B. Oncotic pressure place in the: A. Juxtaglomerular nephrons
C. Renin B. Proximal convoluted tubule
D. Capillary pores C. Cortical nephrons
D. Both A and C
5. Renin is secreted by the nephron in 14. ADH regulates the final urine concentration
response to: A. Low systemic blood by con trolling:
pressure A. Active reabsorption of sodium
B. High systemic blood pressure B. Tubular permeability
C. Oncotic capillary pressure C. Passive reabsorption of urea
D. Increased water retention D. Passive reabsorption of chloride
6. The primary chemical affected by 15. When the body is dehydrated:
the renin angiotensin-aldosterone A. ADH production is decreased
system is: B. ADH production is increased
A. Chloride C. Urine volume is increased
B. Sodium D. Both A and C
C. Potassium 16. Bicarbonate ions filtered by the
D. Hydrogen glomerulus are returned to the blood:
7. Secretion of renin is stimulated by: A. In the proximal convoluted tubule
A. Juxtaglomerular cells B. Combined with hydrogen ions
B. Angiotensin I and II C. By tubular secretion
C. Macula densa cells D. All of the above
D. Circulating angiotensin-converting enzyme 17. If ammonia is not produced by the distal
 convoluted tubule, the urine pH will be:
A. Acidic
B. Basic
18. Place the appropriate letter in front of the
following clearance substances:
A. Exogenous
B. Endogenous
____ inulin
____ creatinine
____ cystatin C
____ 125I-iothalmate
19. The largest source of error in creatinine
clearance tests is:
A. Secretion of creatinine
B. Improperly timed urine specimens
C. Refrigeration of the urine
D. Time of collecting blood sample
20. Given the following information, calculate the
creati nine clearance:
24-hour urine volume: 1000 mL; serum
creatinine: 2.0 mg/dL; urine creatinine: 200
mg/dL
21. Values for creatinine clearance tests on
children are corrected for:
A. Body size
B. Urine volume
C. Activity level
D. Diet
22. Given the data serum creatinine: 1.1 mg/dL;
age: 50 years, and weight: 72 kg, the
estimated creatinine clearance using the
Cockcroft-Gault formula is: A. 46
B. 62
C. 82
D. 127
23. Variables that may be included in estimated
creati nine clearance calculations include all
of the follow ing except:
A. Serum creatinine
B. Urine creatinine
C. Age
D. Blood urea nitrogen
24. An advantage to using cystatin C to monitor
GFR is: A. It does not require urine collection
B. It is not secreted by the tubules
C. It can be measured by immunoassay
D. All of the above
25. Solute dissolved in solvent will:
A. Decrease vapor pressure
B. Lower the boiling point
C. Decrease the osmotic pressure
D. Lower the specific gravity
26. Substances that may interfere with
measurement of urine and serum osmolarity
include all of the follow ing except:
A. Ethanol
B. Lactic acid
C. Sodium
D. Lipids
27. The normal serum osmolarity is:
A. 50–100 mOsm
B. 275–300 mOsm
C. 400–500 mOsm
D. 3 times the urine osmolarity
28. After controlled fluid intake, the
urine-to-serum osmolarity ratio should be
at least:
A. 1:1
B. 2:1
C. 3:1
D. 4:1
©2008 F. A. Davis
29. Calculate the free water clearance from the
following results:
urine volume in 6 hours: 720 mL; urine
osmolarity: 225 mOsm; plasma osmolarity:
300 mOsm
30. To provide an accurate measure of renal
blood flow, a test substance should be
completely:
A. Filtered by the glomerulus
B. Reabsorbed by the tubules
C. Secreted when it reaches the distal convoluted
tubule
D. Cleared on each contact with functional renal tissue
31. Given the following data, calculate the
effective renal plasma flow:
urine volume in 2 hours: 240 mL; urine PAH:
150 mg/dL; plasma PAH: 0.5 mg/dL
32. Renal tubular acidosis can be caused by the:
A. Production of excessively acidic urine due to
increased filtration of hydrogen ions
B. Production of excessively acidic urine due to
increased secretion of hydrogen ions
C. Inability to produce an acidic urine due to
impaired production of ammonia
D. Inability to produce an acidic urine due to
increased production of ammonia
33. Tests performed to detect renal tubular
acidosis after administering an ammonium
chloride load include
all of the following except:
A. Urine ammonia
B. Arterial pH
C. Urine pH
D. Titratable acicity
Case Studies and Clinical
Situations
1. A 44-year-old man diagnosed with acute
 tubular necrosis has a blood urea nitrogen of
60 mg/dL and a blood glucose level of 100
mg/dL. A 2 urine glucose is also reported.
a. State the renal threshold for glucose.
b. What is the significance of the positive
urine glu cose and normal blood
glucose?
2. A patient develops a sudden drop in blood
pressure. a. Diagram the reactions that take
place to ensure
adequate blood pressure within the nephrons.
b. How do these reactions increase blood volume?
c. When blood pressure returns to normal,
how does the kidney respond?
CHAPTER 2 • Renal Function 27
3. A physician would like to prescribe a
nephrotoxic antibiotic for a 60-year-old man
weighing 80 kg. The patient has a serum
creatinine level of 1.0 mg/dL. a. How can the
physician determine whether it is
safe to prescribe this medication before
the patient leaves the office?
b. Can the medication be prescribed to
this patient with a reasonable
assurance of safety?
c. A creatinine clearance was also run on
the patient with the following results: serum
creatinine,
0.9 mg/dL; urine creatinine, 190 mg/dL;
24-hour urine volume, 720 mL. Should
the patient
continue to take the medication?
Justify your answer.
©2008 F. A. Davis
©2008 F. A. Davis
LEARNING OBJECTIVES
Upon completion of this chapter, the reader will be able to:
Introduction
3 R
to
A
Urinalysis
4. A laboratory is obtaining erratic serum osmo
larity results on a patient who is being
monitored at 6 a.m., 12 p.m., 6 p.m., and 12
a.m. Osmolarities are not performed on the
night shift; therefore, the mid night specimen
is run at the same time as the 6 a.m.
specimen.
a. What two reasons could account for
these discrep ancies?
b. If the laboratory is using a freezing point
osmome ter, would these discrepancies
still occur? Why or why not?
c. If a friend was secretly bringing the
patient a pint of whiskey every night,
would this affect the results? Explain
your answer.
5. Following overnight (6 p.m. to 8 a.m.) fluid
depriva tion, the urine-to-serum osmolarity
ratio in a patient who is exhibiting polyuria
and polydipsia is 1:1. The ratio remains the
same when a second specimen is tested at
10 a.m. Vasopressin is then administered
sub cutaneously to the patient, and the fluid
deprivation is continued until 2 p.m., when
another specimen is tested.
a. What disorder do these symptoms and
initial labo ratory results indicate?
b. If the urine-to-serum osmolarity ratio on
the 2 p.m. specimen is 3:1, what is the
underlying cause of the patient’s
disorder?
c. If the urine-to-serum osmolarity ratio on
the 2 p.m. specimen remains 1:1, what is
the underlying cause of the patient’s
disorder?
 recommended urine specimen containers.
5 Describe the correct methodology for
labeling urine specimens.
6 State four possible reasons why a
laboratory would reject a urine
specimen.

1 List three major organic and three major
inor ganic chemical constituents of urine.
2 Describe a method for determining
whether a questionable fluid is urine.
3 Recognize normal and abnormal daily
urine volumes.
4 Describe the characteristics of the
of custody
fasting specimen
first morning specimen
anuria 2-hour postprandial
catheterized specimen chain specimen midstream
clean-catch specimen
KEY TERMS
7 List 10 changes that may take place in a
urine specimen that remains at room
temperature for more than 2 hours.
8 Discuss the actions of bacteria on an
unpreserved urine specimen.
9 Briefly discuss five methods for preserving
urine specimens, including their
advantages and dis - advantages.
10 Instruct a patient in the correct procedure
for collecting a timed urine specimen and
a mid stream clean-catch specimen.
11 Describe the type of specimen needed for
opti mal results when a specific urinalysis
procedure is requested.
nocturia
oliguria
polyuria
suprapubic aspiration
three-glass collection timed
specimen

■ ■ ● History and Importance

Analyzing urine was actually the beginning of
laboratory medicine. References to the study of urine
can be found in the drawings of cavemen and in
Egyptian hieroglyphics, such as the Edwin Smith
Surgical Papyrus. Pictures of early physi cians
commonly showed them examining a bladder-shaped
flask of urine (Fig. 3-1). Often these physicians never
saw the patient, only the patient’s urine. Although these
physi
cians lacked the sophisticated testing mechanisms now
available, they were able to obtain diagnostic
information from such basic observations as color,
turbidity, odor, volume, viscosity, and even sweetness
(by noting that certain speci mens attracted ants).
These same urine characteristics are still reported by
laboratory personnel. However, modern urinaly sis has
expanded beyond physical examination of urine to
include chemical analysis and microscopic examination
of urinary sediment.
Figure 3–1 Physician examines urine flask.
29 (Courtesy of National Library of Medicine)
©2008 F. A. Davis
Many well-known names in the history of
30 CHAPTER 3 • Introduction to Urinalysis medicine are associated with the study of urine,
including Hippocrates, who in the 5th century BC
wrote a book on “uroscopy.” Dur ing the Middle
Ages, physicians concentrated their efforts very
intensively on the art of uroscopy, receiving
instruction in urine examination as part of their
training (Fig. 3-2). By 1140 AD, color charts had
been developed that described the significance of
20 different colors (Fig. 3-3). Chemical testing
progressed from “ant testing” and “taste testing” for
glucose to Frederik Dekkers’ discovery in 1694 of
albuminuria by boiling urine.1
The credibility of urinalysis became
compromised when charlatans without medical
credentials began offering their predictions to the
public for a healthy fee. These charlatans, called
“pisse prophets,” became the subject of a book
pub lished by Thomas Bryant in 1627. The
revelations in this book inspired the passing of the
first medical licensure laws in England—another
contribution of urinalysis to the field of medicine.
The invention of the microscope in the 17th
century led to the examination of urinary sediment
and to the development by Thomas Addis of
methods for quantitating the microscopic sediment.
Richard Bright introduced the concept of urinalysis
as part of a doctor’s routine patient examination in
1827. By the 1930s, however, the number and
complexity of the tests performed in a urinalysis
had reached a point of impracticality, and
urinalysis began to disappear from routine
examinations. Fortunately, devel

Figure 3–2 Instruction in urine examination.

(Courtesy of National Library of Medicine)

opment of modern testing techniques rescued

routine urinalysis, which has remained an integral
part of the patient examination.
Two unique characteristics of a urine specimen
account for this continued popularity:
1. Urine is a readily available and easily
collected specimen.
2. Urine contains information, which can be
obtained by inexpensive laboratory tests,
about many of the body’s major metabolic
functions.
These characteristics fit in well with the current
trends toward preventive medicine and lower
medical costs. In fact, the Clinical and Laboratory
Standards Institute (CLSI) (formerly NCCLS)
defines urinalysis as “the testing of urine with
procedures commonly performed in an expeditious,
reliable, accurate, safe, and cost-effective manner.”
Reasons for performing urinalysis identified by CLSI
include aiding in the diagnosis of disease, screening
asymptomatic populations for undetected disorders,
and monitoring the progress of disease and the
effectiveness of therapy.2
©2008 F. A. Davis

Figure 3–3 A chart used for urine analysis.
(Courtesy of National Library of Medicine)
■ ■ ● Urine Formation
As detailed in Chapter 2, the kidneys
continuously form urine as an ultrafiltrate of
plasma. Reabsorption of water and fil tered
substances essential to body function converts
approxi mately 170,000 mL of filtered plasma to
the average daily urine output of 1200 mL.
■ ■ ● Urine Composition
In general, urine consists of urea and other
organic and inor ganic chemicals dissolved in
water. Urine is normally 95% water and 5%
solutes, although considerable variations in the
concentrations of these solutes can occur owing
to the influ ence of factors such as dietary intake,
physical activity, body metabolism, endocrine
functions, and even body position. Urea, a
metabolic waste product produced in the liver
from the breakdown of protein and amino acids,
accounts for nearly half of the total dissolved
solids in urine. Other organic substances include
primarily creatinine and uric acid. The major
inorganic solid dissolved in urine is chloride,
followed by sodium and potassium. Small or
trace amounts of many additional inorganic
chemicals are also present in urine (Table 3–1).
Dietary intake greatly influences the
concentrations of these inorganic compounds,
making it difficult to establish normal levels. Other
substances found in urine include hormones,
vitamins, and medications. Although not a part of
the original plasma filtrate, the urine also may
contain formed elements, such as cells, casts,
crystals, mucus, and bacteria. Increased
amounts of these formed elements are often indi
cative of disease.
CHAPTER 3 • Introduction to Urinalysis 31
Should it be necessary to determine whether a
particular fluid is urine, the specimen can be tested
for its urea and creatinine content. Because both
these substances are present in much higher
concentrations in urine than in other body fluids, a
high urea and creatinine content can identify a fluid
as urine.
■ ■ ● Urine Volume
Urine volume depends on the amount of water that
the kid neys excrete. Water is a major body
constituent; therefore, the amount excreted is
usually determined by the body’s state of
hydration. Factors that influence urine volume
include fluid intake, fluid loss from nonrenal
sources, variations in the secretion of antidiuretic
hormone, and need to excrete increased amounts
of dissolved solids, such as glucose or salts.
Taking these factors into consideration, although
the normal daily urine output is usually 1200 to
1500 mL, a range of 600 to 2000 mL is considered
normal.
Oliguria, a decrease in urine output, which is
less than 1 mL/kg/hr in infants, less than 0.5
mL/kg/hr in chil dren, and less than 400 mL/day in
adults, is commonly seen when the body enters a
state of dehydration as a result of excessive water
loss from vomiting, diarrhea, perspiration, or severe
burns. Oliguria leading to anuria, cessation of urine
flow, may result from any serious damage to the kid
neys or from a decrease in the flow of blood to the
kid neys. The kidneys excrete two to three times
more urine during the day than during the night. An
increase in the nocturnal excretion of urine is
termed nocturia. Polyuria, an increase in daily
urine volume (greater than 2.5 L/day in adults and
2.5–3 mL/kg/day in children), is often asso ciated
with diabetes mellitus and diabetes insipidus; how
ever, it may be artificially induced by diuretics,
caffeine, or alcohol, all of which suppress the
secretion of antidiuretic hormone.
Diabetes mellitus and diabetes insipidus
produce polyuria for different reasons, and analysis
of the urine is an important step in the differential
diagnosis (Fig. 3-4). Diabetes mellitus is caused by
a defect either in the pancreatic production of
insulin or in the function of insulin, which results in
an increased body glucose concentration. The
kidneys do not reabsorb excess glucose,
necessitating excre tion of increased amounts of
water to remove the dissolved glucose from the
body. Although appearing to be dilute, a urine
specimen from a patient with diabetes mellitus has
a high specific gravity because of the increased
glucose content.
Diabetes insipidus results from a decrease in
the pro duction or function of antidiuretic hormone;
thus, the water necessary for adequate body
 hydration is not reabsorbed from the plasma
filtrate. In this condition, the urine is truly dilute and
has a low specific gravity. Fluid loss in both
diseases is compensated by increased ingestion of
©2008 F. A. Davis
water (polydipsia), producing an even greater
urine volume. Polyuria accompa nied by increased
fluid intake is often the first symptom of either
disease.

32 CHAPTER 3 • Introduction to Urinalysis

Table 3–1 Composition of Urine Collected for

24 Hours Component Amount Remark
Organic
Urea
0.4–1.0 g 0.7 g Benzoic acid is eliminated from the
body in this form; increases with
Creatinine high-vegetable diets
2.9 g
Uric acid Carbohydrates, pigments, fatty
acids, mucin, enzymes, hormones;
may be present in small amounts
Hippuric acid
15.0 g depending on diet and health

Other substances 3.3 g

2.5 g Principal salt; varies with intake

2.5 g Occurs as chloride, sulfate, and

Inorganic
phosphate salts
Sodium chloride (NaCl) Potassium
0.7 g Derived from amino acids
(K )
Occurs primarily as sodium
Sulfate (SO42 ) compounds that serve as buffers in
0.1 g
Phosphate (PO43 ) Ammonium (NH 4 the blood
0.3 g
) 60%–90% of nitrogenous material; Derived from protein metabolism
derived from the metabolism of and glutamine in kidneys; amount
amino acids into ammonia varies depending on blood and
Magnesium (Mg2 ) Calcium (Ca2 ) tissue fluid acidity
Derived from creatine, nitrogenous
25.0–35.0 g Occurs as chloride, sulfate,
substance in muscle tissue
phosphate salts
Common component of kidney
1.5 g stones; derived from catabolism of Occurs as chloride, sulfate,
nucleic acid in food and cell phosphate salts
destruction

Adapted from Tortora, GJ, and Anagnostakos, NP: Principles of Anatomy and Physiology, ed 6, Harper & Row, New York, 1990, p. 51.
■ ■ ● Specimen Collection
As discussed in Chapter 1, urine is a biohazardous
substance that requires the observance of Standard
Precautions. Gloves should be worn at all times when
in contact with the speci men. Specimens must be
collected in clean, dry, leak-proof containers.
Disposable containers are recommended because they
eliminate the chance of contamination due to improper
washing. These disposable containers are available in
a variety of sizes and shapes, including bags with
adhesive for the collection of pediatric specimens and
large containers for 24-hour specimens. Properly
applied screw-top lids are less likely to leak than
snap-on lids.
Containers for routine urinalysis should have a
wide mouth to facilitate collections from female patients
and a wide, flat bottom to prevent overturning. They
should be made of a clear material to allow for
determination of color and clarity. The recommended
capacity of the container is 50 mL, which allows 12 mL
of specimen needed for micro scopic analysis,
additional specimen for repeat analysis, and enough
room for the specimen to be mixed by swirling the
container.
Individually packaged sterile containers with
secure clo sures should be used for microbiologic urine
studies. Sterile
containers are also suggested if more than 2 hours
elapse between specimen collection and analysis.2
All specimens must be labeled properly with the
patient’s name and identification number, the date and
time of collec tion, and additional information such as
the patient’s age and location and the physician’s
name, as required by institutional protocol. Labels must
be attached to the container, not to the lid, and should
not become detached if the container is refrig erated or
frozen.
A requisition form (manual or computerized) must
 accompany specimens delivered to the laboratory. The
infor mation on the form must match the information on
the spec imen label. Additional information on the form Polyuria
can include method of collection or type of specimen,
possible interfering medications, and the patient’s
clinical information. The time the specimen is received Specific gravity
in the laboratory should be recorded on the form.
Improperly labeled and collected specimens
should be rejected by the laboratory, and appropriate CHAPTER 3 • Introduction to Urinalysis 33
personnel should be notified to collect a new specimen.
Unacceptable situations include specimens in place not only in vivo but also in vitro, thus requiring
unlabeled containers, nonmatching labels and correct handling procedures.
requisition forms, specimens contaminated with feces
or toilet paper, containers with contaminated exteriors,
specimens of insufficient quantity, and specimens that
Specimen Integrity
have Following collection, specimens should be delivered to
©2008 F. A. Davis the laboratory promptly and tested within 2 hours. A
specimen that cannot be delivered and tested within 2
hours should be refrigerated or have an appropriate
chemical preservative
Polydipsia
Decreased SG Increased SG changes that may occur in a specimen allowed to
added. Table 3–2 describes the 11 most significant remain unpreserved
under the individ ual test procedures.
Decreased production or
At this point it is important to realize
Function of ADH Increased glucose Diabetes mellitus
that improper preservation can
seriously affect the results of a
routine urinalysis.
Diabetes insipidus at room temperature for longer than
Decreased insulin or 2 hours. Notice that most of the
Decreased function of insulin changes are related to the presence Specimen Preservation
and growth of bacteria.
These variations are discussed again
refrigera tion at 2 C to 8 C, which decreases bacterial
Figure 3–4 Differentiation between diabetes mellitus growth and metabolism. If the urine is to be cultured, it
and diabetes insipidus. should be refrig erated during transit and held
refrigerated until cultured up to 24 hours.2 Refrigeration
can increase the specific gravity, when measured by
been improperly transported. Laboratories should have
urinometer, and the precipitation of amor phous
a written policy detailing their conditions for specimen
phosphates and urates, which may obscure the micro
rejec tion (see Chapter 7).
scopic sediment analysis. The specimen must return to
room temperature before chemical testing by reagent
■ ■ ● Specimen Handling strips. This will correct the specific gravity and may
dissolve some of the amorphous urates.
The fact that a urine specimen is so readily available
When a specimen must be transported over a long
and eas ily collected often leads to laxity in the
dis tance and refrigeration is impossible, chemical
treatment of the spec imen after its collection. Changes
preservatives
in urine composition take
The most routinely used method of preservation is

Table 3–2 Changes in

Unpreserved Urine Analyte Change Cause
Color Urobilinogen
Clarity Decreased
Nitrite
Odor Decreased
Red and white blood cells and casts
pH Decreased
Bacteria
Modified/darkened Decreased Decreased
Glucose Increased Increased
Ketones Increased Decreased
Bilirubin
 Breakdown of urea to ammonia by
Increased urease-producing bacteria/ loss of
Oxidation or reduction of metabolites CO2
Glycolysis and bacterial use
Bacterial growth and precipitation of
Volatilization and bacterial
amorphous material Bacterial
metabolism
multiplication or breakdown of urea
Exposure to light/photo oxidation to
to ammonia
©2008 F. A. Davis
34 CHAPTER 3 • Introduction to Urinalysis
may be added. Commercially prepared transport
tubes are available. The ideal preservative should be
bactericidal, inhibit urease, and preserve formed
elements in the sediment. At the same time, the
preservative should not interfere with chemical tests.
Unfortunately, as can be seen in Table 3–3, the ideal
preservative does not exist; therefore, a preservative
that best suits the needs of the required analysis
should be chosen.
■ ■ ● Types of Specimens
To obtain a specimen that is representative of a
patient’s meta bolic state, regulation of certain
aspects of specimen collec tion is often necessary.
These special conditions may include time, length,
and method of collection and the patient’s dietary and
medicinal intake. It is important to instruct patients
when they must follow special collection procedures.
Frequently encountered specimens are listed in Table
3–4.
Random Specimen
This is the most commonly received specimen
because of its ease of collection and convenience for
the patient. The random specimen may be collected
at any time, but the actual
Table 3–3 Urine
Preservatives Advantages Disadvantages Additional Information
Refrigeration
Thymol
Formalin
Boric acid
(formaldehyde) Toluene
Sodium fluoride
Does not interfere with
chemical tests
biliverdin Oxidation to urobilin
Multiplication of nitrate-reducing
bacteria
Disintegration in dilute alkaline urine
Multiplication
Preservatives
time of voiding should be recorded on the container.2
The random specimen is useful for routine screening
tests to detect obvious abnormalities. However, it may
also show erroneous results resulting from dietary
intake or physical activity just before collection. The
patient will then be requested to collect additional
specimens under more controlled conditions.
First Morning Specimen
Although it may require the patient to make an
additional trip to the laboratory, this is the ideal
screening specimen. It is also essential for preventing
false-negative pregnancy tests and for evaluating
orthostatic proteinuria. The first morning speci men,
or 8-hour specimen, is a concentrated specimen,
thereby assuring detection of chemicals and formed
elements that may not be present in a dilute random
specimen. The patient should be instructed to collect
the specimen immediately on arising and to deliver it to
the laboratory within 2 hours.
Fasting Specimen (Second Morning)
A fasting specimen differs from a first morning
specimen by being the second voided specimen after a
period of fasting.
Preserves glucose and Does not interfere with
sediments well routine tests
Preserves protein and
formed elements well Prevents glycolysis
Does not interfere with rou
tine analyses other than pH
Is a good preservative for
drug analyses
Raises specific gravity by
hydrometer
Excellent sediment preser
Precipitates amorphous
vative
 phosphates and urates tests for glucose, blood, cells and casts
leukocyte esterase, and
Interferes with acid
copper reduction
precipita tion tests for
protein Floats on surface of speci
May precipitate crystals mens and clings to pipettes
Keeps pH at about 6.0
when used in large and testing materials
May use sodium benzoate
amounts Inhibits reagent strip tests Is bacteriostatic (not bacte instead of fluoride for
5
for glucose, blood, and ricidal) at 18 g/L; can use reagent strip testing
leukocytes for culture transport4
Prevents bacterial growth
Interferes with drug and
24 h3
hormone analyses
Rinse specimen container
Acts as a reducing agent, with formalin to preserve
interfering with chemical
©2008 F. A. Davis

CHAPTER 3 • Introduction to Urinalysis 35

Preservatives Advantages Disadvantages Additional Information

routine tests
Phenol Check tablet composition to
Convenient when refrigera May contain one or more of determine possible effects
tion not possible the preservatives including on desired tests
Commercial pre servative
Have controlled concentra sodium fluoride
tablets
tion to minimize
interference
Contains collection cup,
Urine Collection Kits6 C&S preservative tube or
(Becton UA tube
Dickinson, Preservative is boric acid
Rutherford, NJ) Gray C&S Decreases pH; do not use if and may not be used for
Sample stable at room tem urine is below minimum fill UA
tube perature (RT) for 48 hr; line
preserves bacteria
Must refrigerate within Round or conical bottom
Use on automated instru 2 hours
Yellow plain UA tube ments
Bilirubin and urobilinogen Preservative is sodium pro
Cherry red/yellow top tube Stable for 72 hours at RT; may be decreased if pionate; conical bottom
instrument-compatible specimen is exposed to
Saccomanno light and left at RT
Used for cytology studies
Use 1 drop per ounce of
Fixative Preserves cellular elements
specimen
Does not interfere with Causes an odor change
Specimens Type of Specimen Purpose
This specimen will not contain any metabolites from
food ingested before the beginning of the fasting
period. It is recommended for glucose monitoring.7

Table 3–4 Types of Urine

Random 24-h (or timed) Pregnancy tests
Catheterized Orthostatic protein
First morning
Midstream clean-catch Suprapubic Diabetic screening/monitoring

Diabetic monitoring
Fasting (second aspiration
morning)
2-hour postprandial Glucose Optional with blood samples in
glucose tolerance test
Three-glass collection
tolerance test Routine screening Quantitative chemical tests Bacterial

Routine screening
 culture tested for glucose, and the results are a glucose tolerance test (GTT). The
used primarily for monitoring insulin number of specimens varies with the
Routine screening therapy in persons with diabetes length of the test. GTTs may include
Bacterial culture mellitus. A more comprehensive fasting, half hour, 1-hour, 2-hour, and
Bladder urine for bacterial culture evaluation of the patient’s status can 3-hour specimens, and possibly
be obtained if the results of the 4-hour, 5-hour, and 6-hour
Cytology
2-hour postprandial specimen are specimens. The urine is tested for
Prostatic infection compared with those of a fasting glucose and ketones, and the results
specimen and corresponding blood are reported along with the blood test
2-Hour Postprandial glucose tests. results as an aid to interpreting the
Specimen patient’s ability to metabolize a
The patient is instructed to void Glucose Tolerance Specimens measured amount of glucose and are
correlated with the renal threshold for
shortly before consuming a routine
Glucose tolerance specimens are glucose. Collection of these
meal and to collect a specimen 2
sometimes collected to correspond specimens is an institutional option.8
hours after eating. The specimen is
with the blood samples drawn during
©2008 F. A. Davis urine volume produced during that time. Addition of
urine formed before the start of the collec tion period
or failure to include urine produced at the end of the
36 CHAPTER 3 • Introduction to Urinalysis
collection period will produce inaccurate results.
On its arrival in the laboratory, a 24-hour
specimen must be thoroughly mixed and the volume
PROCEDURE 24-Hour (Timed) accurately measured and recorded. If only an aliquot
Specimen Collection is needed for testing, the amount saved must be
Procedure adequate to permit repeat or addi tional testing. If a
specimen is collected in two containers, the contents
Provide patient with written instructions, and of the containers should be combined and thor oughly
explain collection procedure. mixed before aliquoting. Consideration also must be
Issue proper collection container and preservative. given to the preservation of specimens collected over
Day 1: 7 a.m.: patient voids and discards extended periods. All specimens should be
specimen; collects all urine for the next 24 refrigerated or kept on ice during the collection period
hours. and may also require addition of a chemical
Day 2: 7 a.m.: patient voids and adds this urine to preservative. The preservative chosen must be
pre viously collected urine. nontoxic to the patient and should not interfere with
On arrival at laboratory, the entire 24-hour the tests to be performed. Appropriate collection
specimen is thoroughly mixed, and the volume information
is measured and recorded.
An aliquot is saved for testing and additional or
repeat testing; discard remaining urine.

is included with test procedures and should be read

24-Hour (or Timed) Specimen
Measuring the exact amount of a urine chemical is
often necessary instead of just reporting its presence
or absence. A carefully timed specimen must be
used to produce accurate quantitative results. Many
solutes exhibit diurnal variations such as
catecholamines, 17-hydroxysteroids, and electrolytes
in which the lowest concentration is in the early
morning and the highest concentration occurs in the
afternoon.2 When the concentration of the substance
to be measured changes with diurnal variations and
with daily activities such as exercise, meals, and body
metabolism, 24-hour collection is required. If the
concentration of a particular substance remains con
stant, the specimen may be collected over a shorter
period. Care must be taken, however, to keep the
patient adequately hydrated during short collection
periods. Patients must be instructed on the procedure
for collecting a timed specimen.
To obtain an accurate timed specimen, the
patient must begin and end the collection period with
an empty bladder. The concentration of a substance
in a particular period must be calculated from the
before issuing a container and instructions to the
patient.
Catheterized Specimen
This specimen is collected under sterile conditions by
passing a hollow tube (catheter) through the urethra
into the bladder. The most commonly requested test on
a catheterized speci men is a bacterial culture. If a
routine urinalysis is also requested, the culture should
be performed first to prevent contamination of the
specimen.
A less frequently encountered type of catheterized
speci men measures functions in the individual
kidneys. Specimens from the right and left kidneys are
collected separately by passing catheters through the
ureters of the respective kidneys.
Midstream Clean-Catch Specimen
As an alternative to the catheterized specimen, the
midstream clean-catch specimen provides a safer,
less traumatic method for obtaining urine for bacterial
culture and routine urinaly sis. It provides a specimen
 that is less contaminated by epithe lial cells and
bacteria and, therefore, is more representative of the
actual urine than the routinely voided specimen.
Patients must be provided with appropriate cleansing
materi als, a sterile container, and instructions for
cleansing and voiding. Strong bacterial agents, such as
hexachlorophene or povidone-iodine, should not be
used as cleansing agents. Mild antiseptic towelettes
are recommended. Patients are instructed to wash their
hands before beginning the collec tion. Male patients
should clean the glans, which begins at the urethra,
and withdraw the foreskin, if necessary. Female
patients should separate the labia and clean the
urinary mea tus and surrounding area. When
cleansing is complete, patients are to void first into the
toilet, then collect an ade quate amount of urine in the
sterile container, and finish voiding into the toilet. Care
should be taken not to contami nate the specimen
container. As with a catheterized specimen, if a routine
urinalysis is also requested, the culture should be
performed first to prevent contamination of the
specimen.
Suprapubic Aspiration
Occasionally urine may be collected by external
introduction of a needle through the abdomen into the
bladder. Because the bladder is sterile under normal
conditions, suprapubic aspiration provides a sample
for bacterial culture that is com pletely free of
extraneous contamination. The specimen can also be
used for cytologic examination.
Prostatitis Specimen
Similar to the midstream clean-catch collection, the
three glass collection procedure is used to determine
prostatic infection. Instead of discarding the first urine
passed, it is collected in a sterile container. Next, the
midstream portion is collected in another sterile
container. The prostate is then massaged so that
prostate fluid will be passed with the
©2008 F. A. Davis
remaining urine into a third sterile container.
Quantitative cultures are performed on all
specimens, and the first and third specimens are
examined microscopically. In prostatic infection, the
third specimen will have a white blood
cell/high-power field count and a bacterial count 10
times that of the first specimen. Macrophages
containing lipids may also be present. The second
specimen is used as a control for bladder and kidney
infection. If it is positive, the results from the third
specimen are invalid because infected urine has con
taminated the specimen.9 Variations of the procedure
include the Stamey-Mears four-glass localization
method and the pre and postmassage test (PPMT).
The four-glass method consists of bacterial cultures
of the initial voided urine (VB1), mid stream urine
(VB2), expressed prostatic secretions (EPS), and a
postprostatic massage urine specimen (VB3).
Urethral infec tion or inflammation is tested for by the
VB1, and the VB2 is tested for urinary bladder
infection. The prostatic secretions are cultured and
examined for white bood cells. More than 10 to 20
white blood cells per high-power field is considered
abnormal. In the PPMT test, a clean-catch
midstream urine specimen is collected. A second
urine sample is collected after the prostate is
massaged. A positive result is significant bac teriuria
in the postmassage specimen of greater than 10
times the premassage count.10
Pediatric Specimen
Collection of pediatric specimens can present a
challenge. Soft, clear plastic bags with
hypoallergenic skin adhesive to attach to the genital
area of both boys and girls are available for
collecting routine specimens. Sterile specimens may
be obtained by catheterization or by suprapubic
aspiration. Spec imens for culture also may be
obtained using a clean-catch cleansing procedure
and a sterile collection bag. Care must be taken not
to touch the inside of the bag when applying it. For
quantitative testing, bags are available that allow a
tube to be attached and excess urine transferred to
a larger container.
Drug Specimen Collection
Urine specimen collection is the most vulnerable part
of a drug-testing program. Correct collection
procedures and documentation are necessary to
ensure that the results are those of the specific
individual submitting the specimen. The chain of
custody (COC) is the process that provides this doc
umentation of proper sample identification from the
time of collection to the receipt of laboratory results.
The COC is a standardized form that must document
and accompany every step of drug testing, from
collector to courier to laboratory to medical review
officer to employer.
For urine specimens to withstand legal scrutiny,
it is nec essary to prove that no tampering of the
specimen occurred, such as substitution,
adulteration, or dilution. All personnel handling the
specimen must be noted. The specimen must be
handled securely, with a guarantee that no
unauthorized access to the specimen was possible.
Proper identification of the individual whose
information is indicated on the label is
CHAPTER 3 • Introduction to Urinalysis 37
PROCEDURE Urine Drug
Specimen Collection
Procedure11,12
1. The collector washes hands and wears
gloves. 2. The collector adds bluing agent (dye)
to the toilet water reservoir to prevent an
adulterated specimen. 3. The collector
eliminates any source of water other than toilet
 by taping the toilet lid and faucet han dles.
4. The donor provides photo identification or
positive identification from employer
representative.
5. The collector completes step 1 of the
chain of custody (COC) form and has the
donor sign the form.
6. The donor leaves his or her coat, briefcase,
and/or purse outside the collection area to
avoid the possi bility of concealed substances
contaminating the urine.
7. The donor washes his or her hands and
receives a specimen cup.
8. The collector remains in the restroom but
outside the stall, listening for unauthorized
water use, unless a witnessed collection is
requested.
9. The donor hands specimen cup to the
collector. Transfer is documented.
10. The collector checks the urine for abnormal
color and for the required amount (30–45 mL).
11. The collector checks that the temperature strip
on the specimen cup reads 32.5 –37.7 C. The
collector records the in-range temperature on
the COC form (COC step 2). If the specimen
temperature is out of range or the specimen is
suspected to have been diluted or adulterated,
a new specimen must be col lected and a
supervisor notified.
12. The specimen must remain in the sight of the
donor and collector at all times.
13. With the donor watching, the collector peels
off the specimen identification strips from the
COC form (COC step 3) and puts them on the
capped bottle, covering both sides of the cap.
14. The donor initials the specimen bottle seals.
15. The date and time are written on the seals.
16. The donor completes step 4 on the COC
form. 17. The collector completes step 5 on the
COC form. 18. Each time the specimen is
handled, transferred, or
placed in storage, every individual must be
identi fied and the date and purpose of the
change recorded.
19. The collector follows laboratory-specific
instructions for packaging the specimen
bottles and laboratory copies of the COC form.
20. The collector distributes the COC copies to
appro priate personnel.
©2008 F. A. Davis
38 CHAPTER 3 • Introduction to Urinalysis
required. Either photo identification or positive
identification by an employer representative with
photo ID is acceptable. Urine specimen collections
may be “witnessed” or “unwitnessed.” The decision to
obtain a witnessed collection is indicated when it is
suspected that the donor may alter or substitute the
specimen or it is the policy of the client order ing the
test. If a witnessed specimen collection is ordered, a
same-gender collector will observe the collection of
30 to 45 mL of urine. Witnessed and unwitnessed
collections should be immediately handed to the
collector.
The urine temperature must be taken within 4
minutes from the time of collection to confirm the
specimen has not been adulterated. The temperature
should read within the range of 32.5 C to 37.7 C. If
the specimen temperature is not within range, the
temperature should be recorded and the supervisor or
employer contacted immediately. Urine temperatures
outside of the recommended range may indicate
specimen contamination. Recollection of a second
specimen as soon as possible will be necessary. The
urine color is inspected to identify any signs of
contaminants. The specimen is labeled, packaged,
and transported following laboratory specific
instructions.
References
1. Herman, JR: Urology: A View Through the
Retrospectroscope. Harper & Row, Hagerstown, Md.,
1973.
2. Clinical and Laboratory Standards Institute (formerly
NCCLS), Approved Guideline GP16-A2: Urinalysis and
Collection, Transportation, and Preservation of Urine
Specimens; Approved Guideline—Second Edition, CLSI,
formerly NCCLS, Wayne, Pa., 2001.
3. Culhane, JK: Delayed analysis of urine. J Fam Pract
30(4): 473-474, 1990.
4. Meers, PD, and Chow, CK: Bacteriostatic and
bactericidal actions of boric acid against bacteria
and fungi commonly found in urine. J Clin Pathol
43:484-487, 1990.
5. Onstad, J, Hancock, D, and Wolf, P: Inhibitory effect of
fluo ride on glucose tests with glucose oxidase strips.
Clin Chem 21:898-899, 1975.
6. Becton, Dickinson and Company: BD Vacutainer® Urine
Prod ucts for collection, storage, and transport of urine
specimens. Product Circular, 2004.
7. Guthrie, D, Hinnen, D, and Guthrie, R:
Single-voided vs. double-voided urine testing.
Diabetes Care 2(3):269-271, 1979.
8. Baer, DM: Glucose tolerance test: Tips from the clinical
experts. Medical Laboratory Observer, Sept. 2003.
9. Rous, SN: The Prostate Book. Consumers Union, Mt.
Vernon, N.Y., 1988.
10. Stevermer, JJ, and Easley, SK: Treatment of prostatitis.
Am Fam Physician 61(10), 2000.
11. Saint Joseph Hospital Toxicology Laboratory/Creighton
Medical Laboratories, Urine Drug Screening Collection
Procedure, Omaha, Nebr., 1996.
12. STA United, Inc. and the Nebraska Department of
Roads Fed eral Transit Administration Compliance 101
Seminar Work book. Omaha, Nebr., 1996.
STUDY
QUESTIONS
1. The primary chemical constituents of
normal urine are:
A. Protein, sodium, and water
B. Urea, water, and protein
C. Urea, chloride, and water
D. Urea, bilirubin, and glucose
2. An unidentified fluid is received in the laboratory
 with a request to determine if the fluid is urine or 10. What three changes will affect the results of the
another body fluid. Using routine laboratory tests, microscopic examination of urine if it is not tested
what tests within 2 hours?
would determine that the fluid is most probably A. Decreased bacteria, decreased red blood cells,
urine? A. Glucose and ketones decreased casts
B. Urea and creatinine B. Increased bacteria, increased red blood cells,
C. Uric acid and amino acids increased casts
D. Protein and amino acids C. Increased bacteria, decreased red blood cells,
3. A person exhibiting oliguria would have a daily decreased casts
urine volume of: D. Decreased bacteria, increased red blood cells,
A. 200–400 mL increased casts
B. 600–1000 mL 11. What is the method of choice for preservation
C. 1000–1500 mL of rou tine urinalysis samples?
D. Over 1500 mL A. Boric acid
B. Formalin
4. A patient presenting with polyuria, nocturia,
C. Refrigeration
polydip sia, and a high urine specific gravity is
D. Sodium fluoride
exhibiting symptoms of what disorder?
A. Diabetes insipidus 12. For best preservation of urinary sediments, the
B. Diabetes mellitus preservatives of choice are:
C. Urinary tract infection A. Boric acid and thymol
D. Uremia B. Formalin and sodium fluoride
C. Toluene and freezing
5. True or False: Disposable containers with a D. Chloroform and refrigeration
capacity of 50 mL are recommended for the
collection of speci mens for routine urinalysis. 13. What chemical can be used to preserve a
specimen for a culture and a routine
6. The correct method for labeling urine specimen urinalysis?
con tainers is to: A. Boric acid
A. Attach the label to the lid B. Formalin
B. Attach the label to the bottom C. Sodium fluoride
C. Attach the label to the container D. Thymol
D. Use only a wax pencil for labeling
7. A urine specimen for routine urinalysis
would be rejected by the laboratory CHAPTER 3 • Introduction to Urinalysis 39
because:
A. The specimen had been refrigerated
B. More than 50 mL was in the container 14. True or False: A properly labeled urine
C. The specimen and accompanying specimen for routine urinalysis delivered to the
requisition did not match laboratory in a gray-top blood collection tube can
D. The label was placed on the side of the container be tested.
©2008 F. A. Davis
15. What is the specimen of choice for routine
urinalysis? A. Fasting specimen
B. First morning specimen
C. Random specimen
D. 24-Hour specimen
8. An unpreserved specimen collected at 8 a.m.
16. Quantitative urine tests are
and remaining at room temperature until the
performed on: A. First morning
afternoon
specimens
shift arrives can be expected to have:
B. Timed specimens
1. Decreased glucose and ketones
C. Midstsream clean-catch specimens
2. Increased bacteria and nitrite
D. Suprapubic aspirations
3. Decreased pH and turbidity
4. Increased cellular elements 17. Three types of urine specimens that would be
A. 1, 2, and 3 accept able for culture to diagnose a bladder
B. 1, 2, and 4 infection include all of the following except:
C. 1 and 2 only A. Catheterized
D. 4 only B. Midstream clean-catch
9. A specimen containing precipitated amorphous C. Random
urates may have been preserved using: D. Suprapubic aspiration
A. Boric acid 18. A negative urine pregnancy test performed on
B. Chloroform a ran dom specimen may need to be repeated
C. Formalin using a: A. Clean-catch specimen
D. Refrigeration B. Fasting specimen
 C. First morning specimen cells in specimen #1 is significantly lower
D. 24-Hour specimen than in speci men #3, what is the
significance?
19. Cessation of urine flow is termed:
A. Anuria 4. A worker suspects that he or she will be
B. Azotemia requested to collect an unwitnessed urine
C. Diuresis specimen for drug analy sis. He or she carries a
D. Dysuria substitute specimen in his or her pocket for 2
days before being told to collect the spec imen.
20. Persons taking diuretics can be expected to
Shortly after the worker delivers the specimen, he
produce: A. Oliguria
or she is instructed to collect another specimen.
B. Polyuria
a. What test was performed on the specimen to
C. Proteinuria determine possible specimen manipulation? b.
D. Pyuria How was the specimen in this situation
21. What type of urine specimen should be affected? c. If a specimen for drug analysis tests
collected from a patient who complains of positive, state a possible defense related to
painful urination and the physician has specimen collection and handling that an
ordered a routine urinalysis and urine attorney might employ. d. How can this defense
culture? be avoided?
A. Random ©2008 F. A. Davis
B. First morning
C. Fasting
D. Midstream clean-catch
©2008 F. A. Davis

40 CHAPTER 3 • Introduction to Urinalysis

Case Studies and Clinical Situations

1. A 24-hour urine collection received in the
laboratory for creatinine analysis has a volume of
500 mL.
a. Should this specimen be rejected and a new
speci men requested? Why or why not?
b. State a possible source of error, if the creatinine
concentration per 24 hours is abnormally low.
Physical
Examination
2. Mary Johnson brings a urine specimen to the
labora tory for a glucose analysis. The test result
is negative. The physician questions the result
because the patient has a family history of
diabetes mellitus and is experi encing mild
clinical symptoms.
a. What two sources of error related to the urine
specimen could account for the negative test
result?
of Urine
b. How could a specimen be collected that would
more accurately reflect Mary’s glucose metabolism?
3. A three-glass specimen for determination of
possible prostatic infection is sent to the LEARNING OBJECTIVES
laboratory. Specimens #1 and #3 contain Upon completion of this chapter, the reader will be able to:
increased white blood cell levels.

a. If all three specimens have positive bacterial

cul tures, does the patient have a prostatic
infection? Explain your answer.
R

T
4
b. Why is the presence of white blood cells in P

speci men #2 not part of the examination?

A

c. If the amount of bacteria and white blood C


1 List the common terminology used to
report normal urine color.
2 Discuss the relationship of urochrome to
normal urine color.
3 State how the presence of bilirubin in a
specimen may be suspected.
4 Discuss the significance of cloudy red urine
and clear red urine.
5 Name two pathologic causes of black or
brown urine.
6 Discuss the significance of phenazopyridine
in a specimen.
7 State the clinical significance of urine
harmonic oscillation
densitometry
hypersthenuric
hyposthenuric isosthenuric
clarity
©2008 F. A. Davis
42 CHAPTER 4 • Physical Examination of Urine
The physical examination of urine includes the
determination of the urine color, clarity, and specific
gravity. As mentioned in Chapter 3, early physicians
based many medical decisions on the color and clarity
of urine. Today, observation of these characteristics
provides preliminary information concerning disorders
such as glomerular bleeding, liver disease, inborn
errors of metabolism, and urinary tract infection.
Measure ment of specific gravity aids in the
evaluation of renal tubular function. The results of the
physical portion of the urinalysis also can be used to
confirm or to explain findings in the chemical and
clarity. 8 List the common terminology used
to report clarity.
9 Describe the appearance and discuss the
signifi cance of amorphous phosphates and
amorphous urates in freshly voided urine.
KEY TERMS
10 List three pathologic and four
nonpathologic causes of cloudy urine.
11 Define specific gravity, and tell why this
measure ment can be significant in the routine
analysis. 12 Describe the principles of the
urinometer, refrac tometer, and harmonic
oscillation densitometry methods for determining
specific gravity.
13 State two advantages of performing specific
grav ity by refractometer rather than by
urinometer. 14 Given the concentration of
glucose and protein in a specimen, calculate the
correction needed to compensate for these
high-molecular-weight substances in the
refractometer specific gravity reading.
15 Name two nonpathogenic causes of
abnormally high specific gravity readings
using a refrac tometer.
16 State possible causes of abnormal urine odor.
refractometry specific
gravity urinometry
41
microscopic areas of urinalysis.
■ ■ ● Color
The color of urine varies from almost colorless to
black. These variations may be due to normal
metabolic functions, physi cal activity, ingested
materials, or pathologic conditions. A noticeable
change in urine color is often the reason a patient
seeks medical advice; it then becomes the
responsibility of the laboratory to determine whether
this color change is normal or pathologic. The more
common normal and pathologic cor relations of urine
colors are summarized in Table 4–1.

Normal Urine Color
Terminology used to describe the color of normal urine
may differ slightly among laboratories but should be
consistent within each laboratory. Common
descriptions include pale yellow, yellow, dark yellow,
and amber. Care should be taken to examine the
specimen under a good light source, looking down
through the container against a white background. The
yellow color of urine is caused by the presence of a
pigment, which Thudichum named urochrome in 1864.
Urochrome is a product of endogenous metabolism,
and under normal con ditions the body produces it at a
constant rate. The actual amount of urochrome
produced is dependent on the body’s metabolic state,
with increased amounts produced in thyroid conditions
and fasting states.1 Urochrome also increases in urine
that stands at room temperature.2
Because urochrome is excreted at a constant rate,
the intensity of the yellow color in a fresh urine
specimen can give a rough estimate of urine
concentration. A dilute urine will be pale yellow and a
concentrated specimen will be dark yellow. Remember
that, owing to variations in the body’s state of hydration,
these differences in the yellow color of urine can be
normal.

Table 4–1 Laboratory Correlation of Urine

11
Color Color Cause Clinical/Laboratory Correlations
Colorless Yellow foam when shaken and
Acriflavine positive chemical test results for
Pale yellow bilirubin
Phenazopyridine (Pyridium)
Negative bile test results and
possible green fluorescence Drug
Dark yellow Amber Nitrofurantoin commonly administered for urinary
Phenindione tract infections May have orange
Orange foam and thick orange pigment that
Bilirubin oxidized to biliverdin can obscure or interfere with reagent
strip readings
Pseudomonas infection Antibiotic administered for urinary
tract infections
Amitriptyline
Anticoagulant, orange in alkaline
Methocarbamol (Robaxin) Clorets
urine, colorless in acid urine
Indican
Colored foam in acidic urine and
Methylene blue
Yellow-green Yellow-brown false-negative chemical test results
Phenol for bilirubin
Green Commonly observed with random
Blue-green specimens Positive urine culture
Recent fluid consumption Increased 24-hour volume Antidepressant
Polyuria or diabetes insipidus Elevated specific gravity and positive Muscle relaxant, may be
Diabetes mellitus glucose test result Recent fluid green-brown
Dilute random specimen consumption None
Bacterial infections
Concentrated specimen May be normal after strenuous
Fistulas
exercise or in first morning specimen When oxidized

Bilirubin Deydration from fever or burns

©2008 F. A. Davis

CHAPTER 4 • Physical Examination of Urine 43

Color Cause Clinical/Laboratory Correlations

Pink
Red
 screening test or fluorescence under
RBCs oxidized to methemo globin
ultra violet light
Methemoglobin
Alkaline urine of genetically
Homogentisic acid (alkap tonuria) susceptible persons
Melanin or melanogen Tuberculosis medication
Cloudy specimen with RBCs, mucus,
Phenol derivatives and clots
Argyrol (antiseptic)
Seen in acidic urine after standing;
Methyldopa or levodopa
positive chemical test result for blood
Metronidazole (Flagyl)
Brown Black Denatured hemoglobin
RBCs Cloudy urine with positive chemical
Seen in alkaline urine after standing;
test results for blood and RBCs
specific tests are available
visible microscopically
Hemoglobin
Clear urine with positive chemical
Urine darkens on standing and
test results for blood; intravascular
Myoglobin reacts with nitroprusside and ferric
hemolysis
Porphyrins chloride
Clear urine with positive chemical
Interfere with copper reduction tests
test results for blood; muscle
Color disappears with ferric chloride
Beets damage Negative chemical test
Antihypertensive
Rifampin results for blood
Darkens on standing
Menstrual contamination Detect with Watson-Schwartz
azo-gantrisin compounds to persons with urinary tract
infections. This thick, orange pigment not only
Two additional pigments, uroerythrin and obscures the natural color of the specimen but also
urobilin, are also present in the urine in much smaller interferes with chemical tests that are based on color
quantities and con tribute little to the color of normal, reactions. Recognition of the presence of
fresh urine. The presence of uroerythrin, a pink phenazopyridine in a specimen is important so that
pigment, is most evident in specimens that have been laboratories can use alternate testing procedures.
refrigerated, resulting in the precipitation of amorphous Specimens containing phenazopyridine produce a
urates. Uroerythrin attaches to the urates, pro ducing a yellow foam when shaken, which could be mis taken
pink color to the sediment. Urobilin, an oxidation for bilirubin.
product of the normal urinary constituent urobilinogen,
imparts an orange-brown color to urine that is not fresh. Red/Pink/Brown
One of the most common causes of abnormal urine
Abnormal Urine Color color is the presence of blood. Red is the usual color
that blood pro duces in urine, but the color may range
As can be seen in Table 4–1, abnormal urine colors are
from pink to brown, depending on the amount of blood,
as numerous as their causes. Certain colors, however,
the pH of the urine, and the length of contact. Red
are seen more frequently and have a greater clinical
blood cells (RBCs) remaining in an acidic urine for
significance than others.
several hours produce a brown urine due to the
oxidation of hemoglobin to methemoglobin. A fresh
Dark Yellow/Amber/Orange brown urine containing blood may also indicate
Dark yellow or amber urine may not always signify a glomerular bleeding resulting from the conversion of
normal concentrated urine but can be caused by the hemoglobin to methemoglobin.3
presence of the abnormal pigment bilirubin. If bilirubin ©2008 F. A. Davis
is present, it will be detected during the chemical
examination; however, its pres ence is suspected if a
44 CHAPTER 4 • Physical Examination of Urine
yellow foam appears when the specimen is shaken.
Normal urine produces only a small amount of rap idly Besides RBCs, two other substances,
disappearing foam when shaken, and a large amount hemoglobin and myoglobin, produce a red urine
of white foam indicates an increased concentration of and result in a positive chem ical test result for
protein. A urine specimen that contains bilirubin may blood (Fig. 4-1). When RBCs are present, the
also contain hep atitis virus, reinforcing the need to urine is red and cloudy; however, if hemoglobin or
follow standard precau myo globin is present, the specimen is red and
tions. The photo-oxidation of large amounts of excreted clear. Distinguish ing between hemoglobinuria and
uro bilinogen to urobilin also produces a yellow-orange myoglobinuria may be possible by examining the
urine; however, yellow foam does not appear when the patient’s plasma. Hemoglobinuria resulting from
specimen is shaken. Photo-oxidation of bilirubin the in vivo breakdown of RBCs is accompanied by
imparts a yellow-green color to the urine. red plasma. Breakdown of skeletal muscle
Also frequently encountered in the urinalysis produces myo globin. Myoglobin is more rapidly
laboratory is the yellow-orange specimen caused by cleared from the plasma than is hemoglobin and,
the administration of phenazopyridine (Pyridium) or therefore, does not affect the color of the plasma.
 Fresh urine containing myoglobin frequently
exhibits a more reddish-brown color than
hemoglobin. The possibility of hemoglobinuria
being produced from the in vitro lysis of RBCs also
must be considered. Chemical tests to distinguish
between hemoglobin and myoglobin are available
(see Chapter 5).
Urine specimens containing porphyrins also
may appear red resulting from the oxidation of
porphobilinogen to por phyrins. They are often
referred to as having the color of port wine.
Nonpathogenic causes of red urine include
menstrual contamination, ingestion of highly
pigmented foods, and medications. In genetically
susceptible persons, eating fresh beets causes a
red color in alkaline urine.4 Ingestion of black
berries can produce a red color in acidic urine.
Many med ications, including rifampin,
phenolphthalein, phenindione, and phenothiazines,
produce red urine.
Brown/Black
Additional testing is recommended for urine
specimens that turn brown or black on standing
and have negative chemical test results for blood,
inasmuch as they may contain melanin or
homogentisic acid. Melanin is an oxidation product
of the colorless pigment, melanogen, produced in
excess when a malignant melanoma is present.
Homogentisic acid, a metabo lite of phenylalanine,
imparts a black color to alkaline urine from persons
with the inborn-error of metabolism, called
alkaptonuria. These conditions are discussed in
Chapter 9. Medications producing brown/black
urines include levodopa, methyldopa, phenol
derivatives, and metronidazole (Flagyl).
Red urine
Clear Cloudy
Blue/Green
Pathogenic causes of blue/green urine color are
limited to bac terial infections, including urinary tract
infection by Pseudomonas species and intestinal
tract infections resulting in increased urinary
plasma Clear plasma
Hemoglobinuria Myoglobinuria Red
Red blood cells present (Hematuria)
• Use a well-mixed specimen.
• View through a clear container.
Figure 4–1 Differentiation of red urine testing • Determine color and clarity.
chemically positive for blood.
©2008 F. A. Davis
indican. Ingestion of breath deodorizers (Clorets)
can result in a green urine color.5 The medications
methocarbamol (Robaxin), methylene blue, and
amitriptyline (Elavil) may cause blue urine.
Observation of specimen collection bags from
hospital ized patients frequently detects abnormally
colored urine. This may signify a pathologic
condition that requires the urine to stand for a
period of time before color development or the
presence of medications. Phenol derivatives found
in certain intravenous medications produce green
urine on oxidation.6 A purple staining may occur in
catheter bags and is caused by the presence of
indican in the urine or a bacterial infection,
frequently caused by Klebsiella or Providencia
species.7
■ ■ ● Clarity
Clarity is a general term that refers to the
transparency/tur bidity of a urine specimen. In
routine urinalysis, clarity is determined in the same
manner that ancient physicians used: by visually
examining the mixed specimen while holding it in
front of a light source. The specimen should, of
course, be in a clear container. Color and clarity are
routinely determined at the same time. Common
terminology used to report clarity includes clear,
hazy, cloudy, turbid, and milky. As discussed under
the section on urine color, terminology should be
con sistent within a laboratory. A description of
urine clarity reporting is presented in Table 4–2.
Normal Clarity
Freshly voided normal urine is usually clear,
particularly if it is a midstream clean-catch
specimen. Precipitation of amorphous phosphates
and carbonates may cause a white cloudiness.
Nonpathologic Turbidity
The presence of squamous epithelial cells and
mucus, partic ularly in specimens from women, can
result in a hazy but nor mal urine.
PROCEDURE Color and
Clarity Procedure
• View against a white
background. • Maintain adequate
room lighting. • Evaluate a
consistent volume of specimen.
 Vaginal creams

CHAPTER 4 • Physical Examination of Urine 45

Table 4–2 Urine Clarity
Clarity Term Table 4–4 Pathologic Causes
Clear No visible particulates, transparent. of Urine Turbidity
Hazy Few particulates, print easily seen
RBCs
through urine.
WBCs
Cloudy Many particulates, print blurred
Bacteria
through urine.
Yeast
Turbid Print cannot be seen through urine.
Nonsquamous epithelial cells
Milky May precipitate or be clotted.
Abnormal crystals
Lymph fluid
Specimens that are allowed to stand or are Lipids
refrigerated also may develop turbidity that is
nonpathologic. As dis cussed in Chapter 3, improper
preservation of a specimen results in bacterial of urine turbidity can be confirmed by chemical tests
growth; this increases specimen turbidity but is not shown in Table 4–5.
representative of the actual specimen. Clear urine is not always normal. However, with
Refrigerated specimens frequently develop a the increased sensitivity of the routine chemical tests,
thick tur bidity caused by the precipitation of most abnormalities in clear urine will be detected prior
amorphous phosphates, carbonates, and urates. to the microscopic analysis. Current criteria used to
Amorphous phosphates and carbon ates produce a determine the necessity of performing a microscopic
white precipitate in urine with an alkaline pH, examination on all urine specimens include both clarity
whereas amorphous urates produce a precipitate in and chemical tests for RBCs, WBCs, bacteria, and
acidic urine that resembles pink brick dust due to the protein.
presence of uroerythyrin.
Additional nonpathologic causes of urine turbidity
include semen, fecal contamination, radiographic ■ ■ ● Specific Gravity
contrast media, talcum powder, and vaginal creams The ability of the kidneys to selectively reabsorb
(Table 4–3). essential chemicals and water from the glomerular
filtrate is one of the body’s most important functions.
Pathologic Turbidity The intricate process of
The most commonly encountered pathologic causes
of turbidity in a fresh specimen are RBCs, white Table 4–5 Laboratory
blood cells (WBCs), and bacteria caused by infection
or a systemic organ disorder. Other less frequently Correlations in Urine Turbidity 11
encountered causes of patho logic turbidity include
abnormal amounts of nonsquamous epithelial cells, Acidic Urine
yeast, abnormal crystals, lymph fluid, and lipids Amorphous urates
(Table 4–4). Radiographic contrast media
The clarity of a urine specimen certainly provides
Alkaline Urine
a key to the microscopic examination results,
because the amount of turbidity should correspond Amorphous phosphates, carbonates
with the amount of material observed under the Soluble With Heat
microscope. Questionable causes
Amorphous urates, uric acid crystals
Soluble in Dilute Acetic Acid
Table 4–3 Nonpathologic
RBCs
Causes
Amorphous phosphates, carbonates
of Urine Turbidity
Insoluble in Dilute Acetic Acid
Squamous epithelial cells
WBCs
Mucus Bacteria, yeast
Amorphous phosphates, carbonates, urates Spermatozoa
Semen, spermatozoa Soluble in Ether
Fecal contamination
Lipids
Radiographic contrast media Lymphatic fluid, chyle
Talcum powder
©2008 F. A. Davis

46 CHAPTER 4 • Physical Examination of Urine

reabsorption is often the first renal function to become

impaired; therefore, an assessment of the kidney’s ability to
0
reabsorb is a necessary component of the routine urinalysis.
10
This evaluation can be performed by measuring the specific
0
20
gravity of the specimen. Specific gravity also detects possible
dehydration or abnormalities in antidiuretic hormone and
10
30

can be used to determine whether specimen concentration is

20
40

adequate to ensure the accuracy of chemical tests.

30
50
Specific gravity is defined as the density of a solution
40
compared with the density of a similar volume of distilled
50
water at a similar temperature. Because urine is actually water
that contains dissolved chemicals, the specific gravity of urine
is a measure of the density of the dissolved chemicals in the
specimen. As a measure of specimen density, specific gravity
is influenced not only by the number of particles present but
also by their size. Large urea molecules contribute more to the
reading than do the small sodium and chloride molecules.
Therefore, because urea is of less value than sodium and chlo
ride in the evaluation of renal concentrating ability, it also may
be necessary to test the specimen’s osmolarity. This procedure
is discussed in Chapter 2. For purposes of routine urinalysis,
however, the specific gravity provides valuable preliminary
information and can be easily performed by direct methods
using a urinometer (hydrometer) or harmonic oscillation den
sitometry (HOD), and by indirect methods using a refrac
tometer or the chemical reagent strip. This chapter will
discuss the physical methods for determining specific gravity.
The chemical reagent strip method is covered in Chapter amount of urine is poured into a proper-size container
5. and the urinometer is added with a spinning motion.
The scale reading is then taken at the bot tom of the
urine meniscus.
Urinometer
The urinometer reading may also need to be
The urinometer consists of a weighted float attached to corrected for temperature, inasmuch as urinometers
a scale that has been calibrated in terms of urine are calibrated to read 1.000 in distilled water at a
specific gravity. The weighted float displaces a volume particular temperature. The cali bration temperature is
of liquid equal to its weight and has been designed to printed on the instrument and is usu
sink to a level of 1.000 in distilled water. The additional
mass provided by the dissolved sub stances in urine Figure 4–2 Urinometers representing various specific
causes the float to displace a volume of urine smaller gravity readings.
than that of distilled water. The level to which the uri
nometer sinks, as shown in Figure 4-2, represents the ally about 20 C. If the specimen is cold, 0.001 must be
speci men’s mass or specific gravity. sub tracted from the reading for every 3 C that the
Urinometry is less accurate than the other specimen tem perature is below the urinometer
methods cur rently available and is not recommended calibration temperature. Conversely, 0.001 must be
by the Clinical and Laboratory Standards Institute added to the reading for every 3 C that the specimen
(CLSI) formerly the National Committee for Clinical measures above the calibration temperature.
Laboratory Standards (NCCLS).8 A major disadvantage A correction also must be calculated when using
of using a urinometer to measure specific gravity is that either the urinometer or the refractometer if large
it requires a large volume (10 to 15 mL) of spec imen. amounts of glucose or protein are present. Both
The container in which the urinometer is floated must glucose and protein are high molecular-weight
be wide enough to allow it to float without touching the substances that have no relationship to renal
sides and deep enough that it does not rest on the concentrating ability but will increase specimen density.
bottom. When using the urinometer, an adequate There fore, their contribution to the specific gravity is
 subtracted to give a more accurate report of the
kidney’s concentrating abil ity. A gram of protein per
deciliter of urine raises the urine spe cific gravity by
0.003, and 1 g glucose/dL adds 0.004 to the reading.
Consequently, for each gram of protein present, 0.003
must be subtracted from the specific gravity reading,
and 0.004 must be subtracted for each gram of glucose
present.
Example
A specimen containing 1 g/dL of protein and 1
g/dL of glucose has a specific gravity reading
of 1.030. Calculate the corrected reading.
1.030 0.003 (protein) 1.027 0.004 (glucose)
1.023 corrected specific gravity
©2008 F. A. Davis
Refractometer
Refractometry, like urinometry, determines the
concentration of dissolved particles in a
specimen. It does this by measuring refractive
index. Refractive index is a comparison of the
veloc ity of light in air with the velocity of light in a
solution. The concentration of dissolved particles
present in the solution determines the velocity
and angle at which light passes through a
solution. Clinical refractometers make use of
these principles of light by using a prism to direct
1. Put one or two drops of
sample on the prism.
OXX
1.0
50
1.0
40
a specific (mono chromatic) wavelength of
daylight against a manufacturer calibrated
specific gravity scale. The concentration of the
specimen determines the angle at which the light
beam enters the prism. Therefore, the specific
gravity scale is calibrated in terms of the angles
at which light passes through the specimen.
CHAPTER 4 • Physical Examination of Urine 47
The refractometer provides the distinct
advantage of determining specific gravity using a
small volume of specimen (one or two drops).
Temperature corrections are not necessary
because the light beam passes through a
temperature compensating liquid prior to being
directed at the specific gravity scale. Temperature
is compensated between 15 C and 38 C.
Corrections for glucose and protein are still
calculated, although refractometer readings are
less affected by particle density than are
urinometer readings.
When using the refractometer, a drop of urine
is placed on the prism, the instrument is focused at
a good light source, and the reading is taken
directly from the specific grav ity scale. The prism
and its cover should be cleaned after each
specimen is tested. Figure 4-3 illustrates the use of
the refractometer.
3. The sample must spread all over
the prism surface.
2. Close the daylight plate gently.
4. Look at the scale through
the eyepiece.
1.0
30
1.0
20
1.0
10
1.0
 00 34

1. 5

35 1.

5 34

1. 0

35
1.
33
0 5
1.
1. 33
3
Figure 4–3 Steps in the use of the urine U.G. 20˚C nD
specific gravity refractometer. (Courtesy
of NSG Precision Cells, Inc., 195G 5. Read the scale where the boundary line 6. Wipe the sample from the prism clean with
Central Ave., Farmingdale, N.Y., 11735.) intercepts it. a tissue paper and water.
©2008 F. A. Davis specimens required. Results are linear up to a specific
gravity of 1.080.
48 CHAPTER 4 • Physical Examination of Urine
Clinical Correlations
The specific gravity of the plasma filtrate entering the
Calibration
glomerulus is 1.010. The term isosthenuric is used to
screw describe

Measure
Figure 4–4 Calibration of the urine-specific gravity refrac
tometer. (Courtesy of NSG Precision Cells, Inc., 195G
Central Ave., Farmingdale, N.Y., 11735.)

Calibration of the refractometer is performed

using dis tilled water that should read 1.000. If
necessary, the instru ment contains a zero set screw
to adjust the distilled water reading (Fig. 4-4). The
calibration is further checked using 5% NaCl, which as
shown in the refractometer conversion tables should
read 1.022 0.001, or 9% sucrose that should read
1.034 0.001. Urine control samples representing low,
medium, and high concentrations should also be run
at the beginning of each shift. Calibration and control
results are always recorded in the appropriate quality
control records.

Harmonic Oscillation Densitometry In Out

Harmonic oscillation densitometry is based on the Oscillator tube

principle that the frequency of a sound wave entering
a solution changes in proportion to the density of the
solution. Shifts in harmonic oscillation are measured,
and relative density is cal culated. A portion of the
urine sample enters a U-shaped glass tube with an
electromagnetic coil at one end and a motion detector
at the other end. An electric current is applied to the
coil, which causes the sound wave to pass (oscillate)
through the urine sample. Its frequency is altered by
the density of the specimen. A microprocessor at the
other end of the tube measures the change in sound
wave frequency, compensates for temperature
variations, and converts the reading to spe cific gravity
that closely correlates with gravimetric measure ment
(Fig. 4-5).8 All dissolved solutes are measured by this
method, and there is no clarification of cloudy
Figure 4–5 Mass gravity meter used to perform specific
gravity measurement by harmonic oscillation densitometry.
(Courtesy of International Remote Imaging Systems,
Chatsworth, Calif.)
urine with a specific gravity of 1.010. Specimens below
1.010 are hyposthenuric, and those above 1.010 are
hypersthenuric. One would expect urine that has
been concentrated by the kidneys to be
hypersthenuric; however, this is not always true.
Normal random specimens may range from 1.003 to
1.035, depending on the patient’s amount of hydration.
Specimens measuring lower than 1.003 probably are
not urine. Most ran dom specimens fall between 1.015
and 1.025, and any ran dom specimen with a specific
 gravity of 1.023 or higher is generally considered
normal. If a patient exhibits consistently low results,
specimens may be collected under controlled con
ditions as discussed in Chapter 2.
Abnormally high results—over 1.035—are seen in
patients who have recently undergone an intravenous
pyelo gram. This is caused by the excretion of the
injected radi ographic contrast media. Patients who are
receiving dextran
Urine Specific Gravity
Measurements
Method Principle
Urinometry Density
Refractometry Refractive index
Harmonic oscillation Density
densitometry
Reagent strip pKa changes of a polyelectrolyte
©2008 F. A. Davis
or other high-molecular-weight intravenous fluids
(plasma expanders) also produce urine with an
abnormally high spe cific gravity. Once the foreign
substance has been cleared from the body, the
specific gravity returns to normal. In these cir
cumstances, urine concentration can be measured
using the reagent strip chemical test or osmometry
because they are not affected by these
high-molecular-weight substances.9 When the
presence of glucose or protein is the cause of high
results, this is detected in the routine chemical
examination. As dis cussed previously, this can be
corrected for mathematically.
Specimens with specific gravity readings greater
than the refractometer or urinometer scale can be
diluted and retested. If this is necessary, only the
decimal portion of the observed specific gravity is
multiplied by the dilution factor. For exam ple, a
specimen diluted 1:2 with a reading of 1.025 would
have an actual specific gravity of a 1.050.
■ ■ ● Odor
Although it is seldom of clinical significance and is
not a part of the routine urinalysis, urine odor is a
noticeable physical property. Freshly voided urine
has a faint aromatic odor. As the specimen stands,
the odor of ammonia becomes more prominent. The
breakdown of urea is responsible for the char
acteristic ammonia odor. Causes of unusual odors
include bacterial infections, which cause a strong,
unpleasant odor, and diabetic ketones, which
produce a sweet or fruity odor. A serious metabolic
defect results in urine with a strong odor of maple
syrup and is appropriately called maple syrup urine
disease. This and other metabolic disorders with
characteristic urine odors are discussed in Chapter
9. Ingestion of certain foods, including onions, garlic,
and asparagus, can cause an unusual or pungent
urine odor. Studies have shown that although
everyone who eats asparagus produces an odor,
only certain genetically predisposed people can
smell the odor.10 Common causes of urine odors are
summarized in Table 4–6.
Table 4–6 Common Causes
of Urine Odor11
Odor Cause
Aromatic Normal
Foul, ammonia-like Bacterial decomposition,
urinary tract infection
Fruity, sweet Ketones (diabetes mellitus,
starvation, vomiting)
Maple syrup Maple syrup urine disease Mousy
Phenylketonuria
Rancid Tyrosinemia
Sweaty feet Isovaleric acidemia
Cabbage Methionine malabsorption Bleach
Contamination
CHAPTER 4 • Physical Examination of Urine 49
References
1. Drabkin, DL: The normal pigment of urine: The
relationship of urinary pigment output to diet and
metabolism. J Biol Chem 75:443-479, 1927.
2. Ostow, M, and Philo, S: The chief urinary pigment:
The rela tionship between the rate of excretion of the
yellow pigment and the metabolic rate. Am J Med Sci
207:507-512, 1944.
3. Berman, L: When urine is red. JAMA 237:2753-2754,
1977. 4. Reimann, HA: Re: Red urine. JAMA
241(22):2380, 1979. 5. Evans, B: The greening of urine:
Still another “Cloret sign.” N Engl J Med 300(4):202,
1979.
6. Bowling, P, Belliveau, RR, and Butler, TJ: Intravenous
medica tions and green urine. JAMA 246(3):216, 1981.
7. Dealler, SF, et al: Purple urine bags. J Urol
142(3):769-770, 1989.
8. Clinical and Laboratory Standards Institute (Formerly
NCCLS). Urinalysis and Collection, Transportation, and
Preservation of Urine Specimens; Approved Guideline –
Second Edition. NCCLS document GP16-A2, Wayne,
Pa., 2001.
9. Smith, C, Arbogast, C, and Phillips, R: Effect of x-ray
contrast media on results for relative density of urine.
Clin Chem 19(4):730-731, 1983.
10. Mitchell, SC, et al: Odorous urine following asparagus
inges tion in man. Experimenta 43(4):382-383, 1987.
11. Henry, JB, Lauzon, RB, and Schumann, GB: Basic
examina tion of urine. In Henry, JB (ed): Clinical
Diagnosis and Management by Laboratory Methods.
WB Saunders, Philadelphia, 1996.

STUDY
QUESTIONS
1. The concentration of a normal urine specimen
can be estimated by which of the following?
A. Color
B. Clarity
C. Foam
D. Odor
2. The normal yellow color of urine is
produced by: A. Bilirubin
B. Hemoglobin
C. Urobilinogen
D. Urochrome
3. A yellow-brown specimen that produces a yellow
foam when shaken can be suspected of
containing:
A. Bilirubin
B. Carrots
C. Hemoglobin
D. Rhubarb
4. A urine that turns black after standing may
contain: A. Homogentisic acid
B. Melanin
C. Methemoglobin
D. All of the above
Continued on following page
©2008 F. A. Davis
50 CHAPTER 4 • Physical Examination of Urine
Continued
5. Specimens that contain intact RBCs can be
visually distinguished from those that contain
hemoglobin
because:
A. Hemoglobin produces a much brighter red
color B. Hemoglobin produces a cloudy, pink
specimen
C. RBCs produce a cloudy specimen
D. RBCs are quickly converted to hemoglobin
6. After eating beets purchased at the local
farmers’ mar ket, Mrs. Williams notices that her
urine is red, but Mr. William’s urine remains
yellow. The Williamses should:
A. Be concerned because red urine always
indicates the presence of blood
B. Not be concerned because all women
produce red urine after eating beets
C. Be concerned because both of them should
have red urine if beets are the cause
D. Not be concerned because only Mrs.
Williams is genetically susceptible to producing
red urine
from beets
7. Specimens from patients receiving treatment for
uri nary tract infections frequently appear:
A. Clear and red
B. Viscous and orange
C. Dilute and pale yellow
D. Cloudy and red
8. Freshly voided normal urine is usually clear;
however, if it is alkaline, a white turbidity may be
present due to:
A. Amorphous phosphates and carbonates
B. Uroerythrin
C. WBCs
D. Yeast
9. Microscopic examination of a clear urine that pro
duces a pink precipitate after refrigeration will
show: A. Amorphous urates
B. Porphyrins
C. Red blood cells
D. Triple phosphate crystals
10. Under what conditions will a port-wine urine
color be observed in a urine specimen?
A. The patient has eaten Clorets.
B. Melanin is present.
C. Urine contains porphyrins.
D. The patient has a Pseudomonas infection.
11. Which of the following specific gravities would
be most likely to correlate with a dark yellow
urine?
A. 1.005
B. 1.010
C. 1.020
D. 1.030
12. True or False: Urine specific gravity is equally
influ enced by the presence of glucose and
sodium. 13. In what circumstance might a
sediment be slightly warmed prior to microscopic
examination?
A. To hemolyze RBCs
B. To dissolve amorphous urates
C. To increase the specific gravity
D. To correct for temperature in determining the
spe cific gravity
14. A urine specific gravity measured by
refractometer is 1.029, and the temperataure of the
urine is 14 C. The specific gravity should be
reported as:
A. 1.023
B. 1.027
C. 1.029
D. 1.032
15. Refractive index compares:
A. Light velocity in solutions with light velocity
in solids
B. Light velocity in air with light velocity in
solutions C. Light scattering by air with light
 scattering by solutions
D. Light scattering by particles in solution
16. Refractometers are calibrated using:
A. Distilled water and protein
B. Distilled water and blood
C. Distilled water and sodium chloride
D. Distilled water and urea
17. A correlation exists between a specific
gravity of 1.050 and a:
A. 2 glucose
B. 2 protein
C. First morning specimen
D. Radiographic dye infusion
18. An alkaline urine turns black upon standing,
devel ops a cloudy white precipitate, and has a
specific gravity of 1.012. The major concern
about this speci men would be:
A. Color
B. Turbidity
C. Specific gravity
D. All of the above
19. The reading of distilled water by the
refractometer is 1.003. You should:
A. Subtract 1.003 from each specimen
reading B. Add 1.003 to each specimen
reading
C. Use a new refractometer
D. Adjust the set screw
20. A urine specimen with a specific gravity of 1.008
has been diluted 1:5. The actual specific gravity
is: A. 1.008
B. 1.040
C. 1.055
D. 5.040
©2008 F. A. Davis
D. Urinary tract infection
25. The microscopic of a cloudy amber urine is
reported as rare WBCs and epithelial cells. What
does this
suggest?
A. Urinary tract infection
B. Dilute random specimen
C. Precipitated amorphous urates
D. Possible mix-up of specimen and sediment
26. A specimen with a strong ammonia odor and a
heavy white precipitate when it arrives in the
laboratory
may require:
A. Collection of a fresh specimen
B. Centrifugation
C. Dilution for specific gravity
D. Testing under a hood
Case Studies and Clinical Situations
1. A concerned male athlete brings a clear, red
urine specimen to the physician’s office.
a. Would you expect to see RBCs in the microscopic
examination? Why or why not?
b. Name two pathologic causes of a clear, red urine.
Under what conditions do these substances appear
in the urine?
c. The patient reported that the urine appeared
cloudy when he collected it the previous evening,
but it was clear in the morning. Is this possible?
Explain your answer.
d. If the urine is chemically negative for blood, what
questions should the physician ask the patient?

CHAPTER 4 • Physical Examination of Urine 51

21. The method for determining a urine specific

gravity that is based on the principle that the 2. Upon arriving at work, a technologist notices
that a urine specimen left beside the sink by
frequency of a
personnel on the nightshift has a black color.
sound wave entering a solution changes in propor
The initial report describes the specimen as
tion to the density of the solution is:
yellow.
A. Colorimetric
a. Should the technologist be concerned
B. Harmonic oscillation densitometry
about this specimen? Explain your answer.
C. Refractometry
b. If the specimen had an initial pH of 6.0 and
D. Urinometry
now has a pH of 8.0, what is the most
22. A specimen with a specific gravity of 1.005 probable cause of the black color?
would be considered: c. If the specimen has a pH of 6.0 and was
A. Isosthenuric sitting uncapped, what is the most probable
B. Hyposthenuric cause of the black color?
C. Hypersthenuric d. If the original specimen was reported to be
D. Not urine red and to contain RBCs, what is a
23. True or False: Specific gravity is of more possible cause of the black color?
diagnostic value than osmolarity in evaluating
renal concentra 3. While performing a routine urinalysis on a
tion ability. specimen collected from a patient in the urology
24. A strong odor of ammonia in a urine specimen clinic, the tech nician finds a specific gravity
could indicate: reading that exceeds the 1.035 scale on the
A. Ketones refractometer.
B. Normal a. If the urinalysis report has a 1 protein and a
C. Phenylketonuria negative glucose, what is the most probable
 cause of this finding?
b. The technician makes a 1:4 dilution of the
speci men, repeats the specific gravity, and
gets a read ing of 1.015. What is the actual
specific gravity?
c. Using 1 mL of urine, how would the
technician make the above dilution?
d. How could a specific gravity be obtained
from this specimen without diluting it?
4. Mrs. Smith frequently shops at the farmer’s
market near her home. She notices her urine
has a red color and brings a sample to her
physician. The specimen tests negative for
blood.
a. What is a probable cause of Mrs. Smith’s red
©2008 F. A. Davis
©2008 F. A. Davis
Chemical
Examination of
Urine
LEARNING OBJECTIVES
Upon completion of this chapter, the reader will be able to:
1 Describe the proper technique for
performing reagent strip testing.
2 List four causes of premature deterioration of
reagent strips, and tell how to avoid them. 3 List
five quality-control procedures routinely per
formed with reagent strip testing.
4 List two reasons for measuring urinary pH,
and discuss their clinical applications.
5 Discuss the principle of pH testing by
reagent strip.
6 Differentiate between prerenal, renal, and
postre nal proteinuria, and give clinical
examples of each.
urine? b. Mrs. Smith collects a specimen at the
physician’s office. The color is yellow and the
pH is 5.5. Is this consistent with the previous
answer? Why or why not?
5. A urinalysis supervisor requests a new
specimen in each of the following situations.
Support or disagree with the decisions.
a. A green-yellow specimen with negative test
results for glucose and bilirubin.
b. A dark yellow specimen that produces a
large amount of white foam.
c. A cloudy urine with a strong odor of
ammonia. d. A hazy specimen with a
specific gravity greater than 1.035 by
refractometer.
R
5
E
7 Explain the “protein error of indicators,” and
list any sources of interference that may
occur with this method of protein testing.
8 Discuss the sulfosalicylic acid (SSA) test for
urine protein, including interpretation and
sources of interference.
9 Describe the unique solubility characteristics
of Bence Jones protein, and tell how they
can be used to perform a screening test for
the presence of this protein.
10 Discuss microalbuminuria including
significance, reagent strip tests, and their
principles.
11 Explain why glucose that is normally
 reabsorbed in the proximal convoluted 18 Describe the chemical principle of the
tubule may appear in the urine, and state reagent strip method for blood testing, and
the renal threshold levels for glucose. list possible causes of interference.
12 Describe the principle of the glucose 19 Discuss methods used to differentiate
oxidase method of reagent strip testing for between hemoglobinuria and myoglobinuria.
glucose, and name possible causes of
20 Outline the steps in the degradation of
interference with this method.
hemo - globin to bilirubin, urobilinogens,
13 Describe the copper reduction method for
and finally urobilin.
detec tion of urinary reducing substances,
and list pos sible causes of interference. 21 Describe the relationship of urinary bilirubin
and urobilinogen to the diagnosis of bile
14 Interpret matching and nonmatching
duct obstruction, liver disease, and
results between the glucose oxidase
hemolytic disor ders.
and the copper reduction tests for
glucose. 22 Discuss the principle of the reagent strip test
for urinary bilirubin, including possible
15 Name the three “ketone bodies” appearing
sources of error.
in urine and three causes of ketonuria.
23 Discuss the advantages and
16 Discuss the principle of the sodium
disadvantages of performing an Ictotest
nitroprusside reaction, including sensitivity
for detection of urine bilirubin.
and possible causes of interference.
17 Differentiate between hematuria,
hemoglobin uria, and myoglobinuria with Continued on following page
regard to the appearance of urine and
53
serum and clinical significance.
©2008 F. A. Davis

LEARNING OBJECTIVES continued

24 State two reasons for increased urine urobilino
gen and one reason for a decreased urine uro
bilinogen.
25 Describe the Watson-Schwartz test used to differ
entiate among urobilinogen, porphobilinogen,
Ehrlich reactive compounds, and the Hoesch
screening test for porphobilinogen.
26 Discuss the principle of the nitrite-reagent-strip
test for bacteriuria.
27 List five possible causes of a false-negative result
in the reagent-strip test for nitrite.
28 State the principle of the reagent strip test for
leukocytes.
29 Discuss the advantages and sources of error of the
reagent strip test for leukocytes.
30 Explain the principle of the chemical test for specific
gravity.
31 Compare reagent strip testing for urine spe cific gravity
with urinometer and refractometer testing.
32 Correlate physical and chemical urinalysis results.

KEY TERMS
hemoglobinuria ketonuria proteinuria
leukocyturia protein error of indicators
microalbuminuria proteinuria
bacteriuria myoglobinuria renal proteinuria
bilirubin orthostatic proteinuria stercobilinogen
glycosuria postrenal proteinuria prerenal urobilinogen
hematuria
for chemical analysis. Reagent strips currently provide
a simple, rapid means for performing medically
significant chemical analysis of urine, including pH,
protein, glucose, ketones, blood, bilirubin,
urobilinogen, nitrite, leukocytes, and specific gravity.
The two major types of reagent strips are
■ ■ ● Reagent Strips manufactured under the tradenames Multistix
(Siemens Medical Solutions Diagnos tics, Tarrytown,
Routine chemical examination of urine has changed N.Y.) and Chemstrip (Roche Diagnostics, Indianapolis,
dramati cally since the early days of urine testing, Ind.). These products are available with single or
owing to the devel opment of the reagent strip method multiple-testing areas, and the brand and number of
tests used are a matter of laboratory preference.
Certain variations relating to chemical reactions,
sensitivity, specificity, and interfering substances occur
among the products and are dis cussed in the following
sections. Reagent strip brands are also specified by
instrumentation manufacturers.
Reagent strips consist of chemical-impregnated
absorbent pads attached to a plastic strip. A
color-producing chemical reaction takes place when
the absorbent pad comes in contact with urine. The
reactions are interpreted by com paring the color
produced on the pad with a chart supplied by the
manufacturer. Several colors or intensities of a color for
each substance being tested appear on the chart. By
care ful comparison of the colors on the chart and the
strip, a semiquantitative value of trace, 1 , 2 , 3 , or 4
can be
54
reported. An estimate of the milligrams per deciliter
present is available for appropriate testing areas.
Automated reagent strip readers also provide Système
International units.
Reagent Strip Technique
Testing methodology includes dipping the reagent strip
com pletely, but briefly, into a well-mixed specimen,
removing excess urine from the strip by running the
edge of the strip on the container when withdrawing it
from the specimen, wait ing the specified length of time
for reactions to take place, and comparing the colored
reactions against the manufacturer’s chart using a
good light source.
Improper technique can result in errors. Formed
ele ments such as red and white blood cells sink to the
bottom of the specimen and will be undetected in an
unmixed speci men. Allowing the strip to remain in the
urine for an extended period may cause leaching of
reagents from the pads. Like wise, excess urine
remaining on the strip after its removal from the
specimen can produce a runover between chemicals
on adjacent pads, producing distortion of the colors. To
ensure against runover, blotting the edge of the strip on
absorbent paper and holding the strip horizontally while
com paring it with the color chart is recommended. The
amount of time needed for reactions to take place
varies between tests and manufacturers, and ranges
from an immediate reaction for pH to 120 seconds for
leukocytes. For the best semiquan titative results, the
manufacturer’s stated time should be fol
©2008 F. A. Davis
PROCEDURE Reagent Strip
Technique
Dip the reagent strip briefly into a well-mixed
uncen trifuged urine specimen at room
temperature.
Remove excess urine by touching the edge
of the strip to the container as the strip is
withdrawn.
Blot the edge of the strip on a disposable
absorbent pad. Wait the specified amount of
time for the reaction to occur.
Compare the color reaction of the strip pads to the
manufacturer’s color chart in good lighting.
lowed; however, when precise timing cannot be
achieved, manufacturers recommend that
reactions be read between 60 and 120 seconds,
with the leukocyte reaction read at 120 sec onds.
A good light source is, of course, essential for
accurate interpretation of color reactions. The
strip must be held close to the color chart without
actually being placed on the chart. Automated
reagent strip instruments standardize the color
interpretation and timing of the reaction and are
not subject to room lighting deficiencies or
inconsistency among labora tory personnel
(Appendix A). Reagent strips and color charts
from different manufacturers are not
interchangeable. Speci mens that have been
refrigerated must be allowed to return to room
temperature prior to reagent strip testing, as the
enzy matic reactions on the strips are
temperature dependent.
Handling and Storage of Reagent Strips
In addition to using correct testing technique,
reagent strips must be protected from
deterioration caused by moisture, volatile
chemicals, heat, and light. Reagent strips are
pack aged in opaque containers with a desiccant
to protect them from light and moisture. Strips are
removed just prior to test ing, and the bottle is
tightly resealed immediately. Bottles should not
be opened in the presence of volatile fumes. Man
ufacturers recommend that reagent strips be
stored at room temperature below 30 C. All
bottles are stamped with an expiration date that
represents the functional life expectancy of the
chemical pads. Reagent strips must not be used
past the expiration date. Care must be taken not
to touch the chemi cal pads when removing the
strips.
Quality Control of Reagent Strips
Reagent strips must be checked with both
positive and nega tive controls a minimum of
once every 24 hours. Many labo ratories perform
this check at the beginning of each shift. Testing
is also performed when a new bottle of reagent
strips is opened, questionable results are
obtained, or there is con cern about the integrity
of the strips. All quality control results must be
recorded following laboratory protocol. Several
com
CHAPTER 5 • Chemical Examination of Urine 55
 ©2008 F. A. Davis
Summary of Reagent Strip Testing
Care of Reagent Strips
1. Store with desiccant in an opaque, tightly
closed con tainer.
2. Store below 30 C; do not freeze.
3. Do not expose to volatile fumes.
4. Do not use past the expiration date.
5. Do not use if chemical pads become
discolored. 6. Remove strips immediately
prior to use.
Technique
1. Mix specimen well.
2. Let refrigerated specimens warm to room
temperature before testing.
3. Dip the strip completely, but briefly, into
specimen. 4. Remove excess urine by withdrawing
the strip against the rim of the container and by
blotting the edge of the strip.
5. Compare reaction colors with the
manufacturer’s chart under a good light source at
the specified time. 6. Perform backup tests when
indicated.
7. Be alert for the presence of interfering
substances. 8. Understand the principles and
significance of the test, read package inserts.
9. Relate chemical findings to each other and to the
physi cal and microscopic urinalysis results.
Quality Control
1.Test open bottles of reagent strips with known
positive and negative controls every 24 hr.
2. Resolve control results that are out of range by
further testing.
3.Test reagents used in backup tests with positive
and neg ative controls.
4. Perform positive and negative controls on new
reagents and newly opened bottles of reagent
strips.
5. Record all control results and reagent lot numbers.
panies manufacture both positive and negative
controls. Dis tilled water is not recommended as a
negative control because reagent strip chemical
reactions are designed to perform at ionic
concentrations similar to urine.1 All readings of the
neg ative control must be negative, and positive
control readings should agree with the published
value by one color block. Results that do not
agree with the published values must be resolved
through the testing of additional strips and controls
(see Chapter 7).
Demonstration of chemically acceptable
reagent strips does not entirely rule out the
possibility of inaccurate results. Interfering
substances in the urine, technical carelessness,
and color blindness also produce errors. Reagent
strip manufac turers have published information
concerning the limitations of their chemical
reactions, and laboratory personnel should
56 CHAPTER 5 • Chemical Examination of Urine
be aware of these conditions. As mentioned in
Chapter 4, a primary example of reagent strip
interference is the masking of color reactions by
the orange pigment present in the urine of persons
taking phenazopyridine compounds. If laboratory
personnel do not recognize the presence of this
pigment or other pigments, they will report many
erroneous results.
Nonreagent strip testing procedures using
tablets and liquid chemicals are available when
questionable results are obtained or highly
pigmented specimens are encountered. In the
past, many of these procedures were used
routinely to confirm positive results. Increased
specificity and sensitivity of reagent strips and the
use of automated strip readers have reduced the
need for routine use of these procedures.2 The
chemical reliability of these procedures also must
be checked using positive and negative controls.
Specific backup tests are discussed in this chapter
under the sections devoted to the chemical
parameters for which they are used.
■ ■ ● pH
Along with the lungs, the kidneys are the major
regulators of the acid-base content in the body.
They do this through the secretion of hydrogen in
the form of ammonium ions, hydro gen phosphate,
and weak organic acids, and by the reabsorp tion
of bicarbonate from the filtrate in the convoluted
tubules (see Chapter 2). A healthy individual
usually produces a first morning specimen with a
slightly acidic pH of 5.0 to 6.0; a more alkaline pH
is found following meals (alkaline tide). The pH of
normal random samples can range from 4.5 to 8.0.
Consequently, no normal values are assigned to
urinary pH, and it must be considered in
conjunction with other patient information, such as
the acid-base content of the blood, the patient’s
renal function, the presence of a urinary tract infec
tion, the patient’s dietary intake, and the age of the
specimen (Table 5–1).
Clinical Significance
The importance of urinary pH is primarily as an aid
in determining the existence of systemic acid-base
disorders of metabolic or respiratory origin and in
the management of urinary conditions that require
the urine to be maintained at a specific pH. In
respiratory or metabolic acidosis not related to
renal function disorders, the urine is acidic;
conversely, if respiratory or metabolic alkalosis is
present, the urine is alkaline. Therefore, a urinary
pH that does not conform to this pattern may be
used to rule out the suspected condi tion, or, as
discussed in Chapter 2, it may indicate a disorder
resulting from the kidneys’ inability to secrete or to
reabsorb acid or base.
 The precipitation of inorganic chemicals
dissolved in the urine forms urinary crystals and
renal calculi. This precipita tion depends on urinary
pH and can be controlled by main taining the urine
at a pH that is incompatible with the precipitation of
the particular chemicals causing the calculi
formation. For example, calcium oxalate, a
frequent con stituent of renal calculi, precipitates
primarily in acidic and
Table 5–1 Causes of Acid
and Alkaline Urine
Acid Urine Alkaline Urine Emphysema
Hyperventilation Diabetes mellitus
Vomiting
Starvation Renal tubular acidosis
Dehydration Presence of urease
producing
bacteria
Diarrhea Vegetarian diet Presence of
acid- Old specimens producing
bacteria
(Escherichia coli)
High-protein diet
Cranberry juice
Medications (methenamine
mandelate [Mandelamine],
fosfomycin tromethamine)
not alkaline urine. Therefore, maintaining urine at
an alkaline pH discourages formation of the calculi.
Knowledge of uri nary pH is important in the
identification of crystals observed during
microscopic examination of the urine sediment. This
will be discussed in detail in Chapter 6.
The maintenance of an acidic urine can be of
value in the treatment of urinary tract infections
caused by urea-splitting organisms because they
do not multiply as readily in an acidic medium.
These same organisms are also responsible for the
highly alkaline pH found in specimens that have
been allowed to sit unpreserved for extended
periods. Urinary pH is controlled primarily by dietary
regulation, although med ications also may be
used. Persons on high-protein and high meat diets
tend to produce acidic urine, whereas urine from
vegetarians is more alkaline, owing to the formation
of bicar bonate following digestion of many fruits
and vegetables. An exception to the rule is
cranberry juice, which produces an acidic urine and
has long been used as a home remedy for minor
bladder infections. Medications prescribed for
urinary tract infections, such as methenamine
mandelate (Mande lamine) and fosfomycin
tromethamine, are metabolized to produce an acidic
urine.
The pH of freshly excreted urine does not
reach 9 in normal or abnormal conditions. A pH of 9
is associated with an improperly preserved
specimen and indicates that a fresh specimen
should be obtained to ensure the validity of the
analysis.
Reagent Strip Reactions
The Multistix and Chemstrip brands of reagent
strips measure urine pH in 0.5- or 1-unit increments
between pH 5 and 9.
©2008 F. A. Davis
Summary of Clinical
Significance
of Urine pH
1. Respiratory or metabolic acidosis/ketosis
2. Respiratory or metabolic alkalosis
3. Defects in renal tubular secretion and
reabsorption of acids and bases—renal tubular
acidosis
4. Renal calculi formation
5.Treatment of urinary tract infections
6. Precipitation/identification of crystals
7. Determination of unsatisfactory specimens
To differentiate pH units throughout this wide
range, both manufacturers use a double-indicator
system of methyl red and bromthymol blue.
Methyl red produces a color change from red to
yellow in the pH range 4 to 6, and bromthymol
blue turns from yellow to blue in the range of 6 to
9. There fore, in the pH range 5 to 9 measured by
the reagent strips, one sees colors progressing
from orange at pH 5 through yel low and green to
a final deep blue at pH 9.
Methyl red H → Bromthymol blue H
(Red-Orange → Yellow) (Green → Blue)
No known substances interfere with urinary pH
measure ments performed by reagent strips.
Care must be taken, how ever, to prevent runover
between the pH testing area and the adjacent,
highly acidic protein testing area on Multistix, as
this may produce a falsely acidic reading in an
alkaline urine.
■ ■ ● Protein
Of the routine chemical tests performed on urine,
the most indicative of renal disease is the protein
determination. The presence of proteinuria is
often associated with early renal disease, making
the urinary protein test an important part of any
physical examination. Normal urine contains very
 little protein: usually, less than 10 mg/dL or 100
mg per 24 hours is excreted. This protein
consists primarily of low-molecular
Summary of pH Reagent Strip
CHAPTER 5 • Chemical Examination of Urine 57
weight serum proteins that have been filtered by
the glomeru lus and proteins produced in the
genitourinary tract. Owing to its low molecular
weight, albumin is the major serum pro tein found
in normal urine. Even though it is present in high
concentrations in the plasma, the normal urinary
albumin content is low because the majority of
albumin presented to the glomerulus is not filtered,
and much of the filtered albu min is reabsorbed by
the tubules. Other proteins include small amounts
of serum and tubular microglobulins, Tamm Horsfall
protein produced by the tubules, and proteins from
prostatic, seminal, and vaginal secretions.
Clinical Significance
Demonstration of proteinuria in a routine analysis
does not always signify renal disease; however, its
presence does require additional testing to
determine whether the protein represents a normal
or a pathologic condition. Clinical proteinuria is
indicated at ≥30 mg/dL (300 mg/L).3 The causes of
Reagents specimens
Sensitivity Nitrite
Leukocytes
Sources of error/ interference Microscopic
the tubular reabsorption capacity and C and 60 C and clear at 100 C can be
is excreted in the urine. When Bence suspected of containing Bence Jones
Correlations with other tests Jones protein is suspected, a
Methyl red, bromthymol blue 5–9 screening test that uses
solubility characteristics of the protein by filtering the specimen at 100 C and
can be performed. Unlike other
No known interfering substances
proteins, which coagulate and remain as it cools to between 40 C and 60 C.
Runover from adjacent pads Old
coagulated when exposed to heat,
©2008 F. A. Davis
58 CHAPTER 5 • Chemical Examination of Urine
sons with multiple myeloma excrete detectable levels
of Bence Jones protein. Suspected cases of multiple
myeloma must be diagnosed by performing serum
electrophoresis and immu noelectrophoresis.
Renal Proteinuria
Proteinuria associated with true renal disease may be
the result of either glomerular or tubular damage.
Glomerular Proteinuria
When the glomerular membrane is damaged,
selective filtra tion is impaired, and increased amounts
proteinuria are varied and can be grouped into
three major categories: pre renal, renal, and
postrenal, based on the origin of the protein.
Prerenal Proteinuria
As the name implies, prerenal proteinuria is caused
by condi tions affecting the plasma prior to its
reaching the kidney and, therefore, is not indicative
of actual renal disease. This condi tion is frequently
transient, caused by increased levels of low
molecular-weight plasma proteins such as
hemoglobin, myoglobin, and the acute phase
reactants associated with infection and
inflammation. The increased filtration of these
proteins exceeds the normal reabsorptive capacity
of the renal tubules, resulting in an overflow of the
proteins into the urine. Because reagent strips
detect primarily albumin, prerenal pro teinuria is
usually not discovered in a routine urinalysis.
Bence Jones Protein
A primary example of proteinuria due to increased
serum pro tein levels is the excretion of Bence
Jones protein by persons with multiple myeloma. In
multiple myeloma, a proliferative disorder of the
immunoglobulin-producing plasma cells, the serum
contains markedly elevated levels of monoclonal
immunoglobulin light chains (Bence Jones protein).
This low molecular-weight protein is filtered in
quantities exceeding
Bence Jones protein coagulates at
temperatures between 40 C and 60 C
and dis solves when the temperature
reaches 100 C. Therefore, a spec
imen that appears turbid between 40
protein. Interference due to other
the unique precipitated proteins can be removed
observing the specimen for turbidity
Not all per
of serum protein and eventually red and white blood
cells pass through the mem brane and are excreted in
the urine. Conditions that present the glomerular
membrane with abnormal substances (e.g., amyloid
material, toxic substances, and the immune com
plexes found in lupus erythematosus and
streptococcal glomerulonephritis) are major causes of
proteinuria due to glomerular damage.
Increased pressure from the blood entering the
glomeru lus may override the selective filtration of the
glomerulus, causing increased albumin to enter the
filtrate. This condition may be reversible, such as
occurs during strenuous exercise and dehydration or
associated with hypertension. Proteinuria that occurs
during the latter months of pregnancy may indi cate a
pre-eclamptic state and should be considered in con
junction with other clinical symptoms, such as
hypertension, to determine if this condition exists.

Tubular Proteinuria
Increased albumin is also present in disorders
affecting tubu lar reabsorption because the normally
filtered albumin can no longer be reabsorbed. Other
low-molecular-weight proteins that are usually
reabsorbed are also present. Causes of tubu lar
dysfunction include exposure to toxic substances and
heavy metals, severe viral infections, and Fanconi
syndrome. The amount of protein that appears in the
urine following glomerular damage ranges from
slightly above normal to 4 g/day, whereas markedly
elevated protein levels are seldom seen in tubular
disorders.
The discovery of protein, particularly in a random
sam ple, is not always of pathologic significance,
because several benign causes of renal proteinuria
exist. Benign proteinuria is usually transient and can
be produced by conditions such as strenuous
exercise, high fever, dehydration, and expo sure to
cold.
Orthostatic (Postural) Proteinuria
A persistent benign proteinuria occurs frequently in
young adults and is termed orthostatic, or postural,
proteinuria. It occurs following periods spent in a
vertical posture and disap pears when a horizontal
position is assumed. Increased pres sure on the renal
vein when in the vertical position is believed
©2008 F. A. Davis
to account for this condition. Patients suspected of
orthostatic proteinuria are requested to empty their
bladder before going to bed, collect a specimen
immediately upon arising in the morning, and collect a
second specimen after remaining in a vertical position
for several hours. Both specimens are tested for
protein, and if orthostatic proteinuria is present, a
negative reading will be seen on the first morning
specimen, and a pos itive result will be found on the
second specimen.
Microalbuminuria
The development of diabetic nephropathy leading to
reduced glomerular filtration and eventual renal failure
is a common occurrence in persons with both type 1
and type 2 diabetes mellitus. Onset of renal
complications can first be predicted by detection of
microalbuminuria, and the progression of renal
disease can be prevented through better stabilization of
blood glucose levels and controlling of hypertension.
The presence of microalbuminuria is also associated
with an increased risk of cardiovascular disease.4,5
Prior to the development of current reagent strip
meth ods that are specific for albumin, detection of
microalbumin uria required collection of a 24-hr urine
specimen. Specimens were tested using quantitative
procedures for albumin. Results were reported in mg of
albumin/24 hours or as the albumin excretion (AER) in
g/min. With these methods, microalbumin is considered
significant when 30 to 300 mg of albumin is excreted in
24 hours or the AER is 20-200 g/min.
Postrenal Proteinuria
Protein can be added to a urine specimen as it passes
through the structures of the lower urinary tract
(ureters, bladder, urethra, prostate, and vagina).
Bacterial and fungal infections and inflammations
produce exudates containing protein from the interstitial
fluid. The presence of blood as the result of injury or
menstrual contamination contributes protein, as does
the presence of prostatic fluid and large amounts of
spermatozoa.
Reagent Strip Reactions
Traditional reagent strip testing for protein uses the
principle of the protein error of indicators to produce
a visible colori metric reaction. Contrary to the general
belief that indicators produce specific colors in
response to particular pH levels, certain indicators
change color in the presence of protein even though
the pH of the medium remains constant. This is
because protein (primarily albumin) accepts hydrogen
ions from the indicator. The test is more sensitive to
albumin because albumin contains more amino groups
to accept the hydrogen ions than other proteins.
Depending on the manu facturer, the protein area of
the strip contains either tetra bromphenol blue or 3′, 3′′,
5′, 5′′-tetrachlorophenol-3, 4, 5,
6-tetrabromosulfonphthalein and an acid buffer to
maintain the pH at a constant level. At a pH level of 3,
both indicators appear yellow in the absence of protein;
however, as the pro
Summary of Clinical Significance
of Urine Protein
Prerenal Tubular Disorders
Intravascular hemolysis Fanconi syndrome
Muscle injury Toxic agents/heavy metals
Acute phase reactants Severe viral infections
Multiple myeloma
Renal Postrenal
Glomerular disorders Lower urinary tract infections/
inflammation
Immune complex Injury/trauma
disorders
Menstrual contamination
Amyloidosis Prostatic fluid/spermatozoa
 readings.
Toxic agents Vaginal secretions
Diabetic nephropathy Sulfosalicylic Acid Precipitation Test
Strenuous exercise
Dehydration The SSA test is a cold precipitation test that reacts
Hypertension equally with all forms of protein (Table 5–2). Various
Pre-eclampsia concentrations
Orthostatic or postural proteinuria

tein concentration increases, the color progresses Summary of Protein Reagent Strip
through various shades of green and finally to blue.
Readings are reported in terms of negative, trace, 1 , Reagents Multistix:Tetrabromphenol blue
2 , 3 , and 4 ; or the semiquantitative values of 30, Chemstrip: 3′, 3′′, 5′, 5′′ tetra
100, 300, or 2000 mg/dL corresponding to each chlorophenol
color change. Trace values are consid ered to be 3, 4, 5, 6-tetrabromosul
less than 30 mg/dL. Interpretation of trace readings fophthalein
can be difficult. Reporting of trace values may be a
laboratory option. The specific gravity of the Sensitivity Multistix: 15–30 mg/dL albumin
specimen should be con sidered because a trace Chemstrip: 6 mg/dL albumin
protein in a dilute specimen is more significant than
in a concentrated specimen. Sources of error/ False-positive: Highly buffered
interference alkaline urine Pigmented specimens,
pH 3.0 phenazopyridine
Indicator Protein → Protein H
(Yellow) Indicator H Quaternary ammonium
compounds (deter
(Blue-green)
gents)
Reaction Interference Antiseptics, chlorhexi
dine
The major source of error with reagent strips occurs Loss of buffer from
with highly buffered alkaline urine that overrides the prolonged exposure
acid buffer system, producing a rise in pH and a of the reagent strip to
color change unrelated to protein concentration. the specimen
Likewise, a technical error of allow ing the reagent High specific gravity
pad to remain in contact with the urine for a False-negative: Proteins other than
prolonged period may remove the buffer. albumin
False-positive read Microalbuminuria

Correlations with Blood

CHAPTER 5 • Chemical Examination of Urine 59 other tests Nitrite
Leukocytes
ings are obtained when the reaction does not take Microscopic
place under acidic conditions. Highly pigmented urine ©2008 F. A. Davis
and contamination of the container with quaternary
ammonium compounds, detergents, and antiseptics 60 CHAPTER 5 • Chemical Examination of Urine
also cause false-positive readings. A false-positive
trace reading may occur in specimens with a high
specific gravity. The fact that reagent strips detect pri PROCEDURE
marily albumin can result in a false-negative reading in
the presence of proteins other than albumin. Sulfosalicylic Acid (SSA) Precipitation Test
Traditionally, most laboratories chose to confirm
all pos itive protein results using the sulfosalicyclic
acid (SSA) pre cipitation test. This practice is being
replaced by more selective criteria to determine the
need for additional testing. For example, some
laboratories perform SSA testing only on highly Table 5–2 Reporting SSA
alkaline urines, and others acidify the specimen and
retest using a reagent strip. Also, a laboratory with an Turbidity Grade Turbidity Protein Range
auto mated strip reader can opt not to record trace (mg/dL)
Grade the degree of Testing Micral-Test
• Add 3 mL of 3% SSA turbidity (see Table 5–2).
reagent to 3 mL of
Microalbumin Summary Principle: Enzyme
centrifuged urine. immunoassay
• Mix by inversion and Immunologic Tests Sensitivity: 0–10 mg/dL
observe for cloudiness. • Reagents: Gold-labeled
 antibody
B-galactosidase
Chlorophenol red
2
galactoside
Interference: False
negative: Dilute urine
Immunodip 3
Negative Trace
1 4 Clumps of protein
No increase in turbidity6
Principle: Immunochromographics
Sensitivity: 1.2–8.0 mg/dL
Reagents: Antibody coated blue latex
particles Interference: False
negative-dilute urine
Albumin: Creatinine Ratio
Clinitest Microalbumin Strips/Multistix-Pro
Principle: Sensitive albumin tests related to
creatinine concentration to correct for patient
hydration
Reagents:
Albumin: diodo-dihydroxydinitrophenyl
tetrabro mosulfonphtalein
Creatinine: copper sulfate,
tetramethylbenzidine,
diisopropylbenzenedihydroperoxide
Sensitivity:
Albumin: 10–150 mg/L
Creatinine: 10–300 mg/dL, 0.9–26.5 mmol/L
Interference:
Visibly bloody or abnormally colored urine
Creatinine: Cimetidine-False Positive
and amounts of SSA can be used to precipitate protein,
and methods vary greatly among laboratories. All
precipitation tests must be performed on centrifuged
specimens to remove any extraneous contamination.
Of course, any substance precipitated by acid ©2008 F. A. Davis
produces false turbidity in the SSA test. The most
frequently encoun tered substances are radiographic
dyes, tolbutamide metabo lites, cephalosporins,
penicillins, and sulfonamides.6 The presence of
radiographic material can be suspected when a
markedly elevated specific gravity is obtained. In the
presence of radiographic dye, the turbidity also
increases on standing due to the precipitation of
crystals rather than protein. The patient’s history
provides the necessary information on tolbu tamide and
antibiotic ingestion. In contrast to the reagent
strip test, a highly alkaline urine produces
false-negative read ings in precipitation tests, as the
higher pH interferes with precipitation. Use of a more
concentrated solution of SSA may overcome the effect
of a highly buffered, alkaline urine.
Testing for Microalbuminuria
Several semiquantitative reagent strip methods have
been developed so that patients at risk for renal
disease can be monitored using random or first
morning specimens. These methods are based on
Noticeable tur bidity
6–30
Distinct turbidity with no
granula tion
30–100 100–200
Turbidity with granulation
with no flocculation
Turbidity with granulation
200–400 400
and flocculation
immunochemical assays for albumin or
albumin-specific reagent strips that also measure
creatinine to produce an albumin:creatinine ratio.
Immunochemical assays include the Micral-Test
(Roche Diagnostics, Indianapolis, Ind.) and the
Immunodip (Diag nostic Chemicals Limited, Oxford,
Canada). Both reagent strips are read visually, and first
morning specimens are rec ommended.
Micral-Test reagent strips contain a gold-labeled
antihu man albumin antibody-enzyme conjugate. Strips
are dipped into the urine up to a level marked on the
strip and held for 5 seconds. Albumin in the urine binds
to the antibody. The bound and unbound conjugates
move up the strip by wick ing action. Unbound
conjugates are removed in a captive zone by
combining with albumin embedded in the strip. The
urine albumin–bound conjugates continue up the strip
and reach an area containing enzyme substrate. The
conjugated enzyme reacts with the substrate,
producing colors ranging from white to red. The amount
of color produced represents the amount of albumin
present in the urine. The color is com pared with a
chart on the reagent strip bottle after 1 minute. Results
range from 0 to 10 mg/dL.
The Immunodip reagent strip uses an
immunochromo graphic technique. Strips are
individually packaged in spe cially designed containers.
The container is placed in the urine specimen for 3
minutes. A controlled amount of urine
enters the container through a vent hole. The
urine encoun ters blue latex particles coated with
antihuman albumin anti body. Albumin in the
urine binds with the coated particles. The bound
and unbound particles continue to migrate up the
strip. The migration is controlled by the size of
the particles; unbound particles do not migrate as
far as the bound parti cles. First a blue band is
formed by the unbound particles. The bound
particles continue to migrate and form a second
blue band further up the strip. The top band
therefore repre sents the bound particles (urine
albumin) and the bottom band represents
unbound particles. The color intensity of the
bands is compared against the manufacturer’s
color chart. A darker bottom band represents
greater than 1.2 mg/dL, equal band colors
represent 1.2 to 1.8 mg/dL, and a darker top
band represents 2.0 to 8.0 mg/dL of albumin. A
darker bot tom band is negative, equal band
 color is borderline, and a darker top band
represents positive results.
Albumin: Creatinine Ratio
The Clinitek Microalbumin reagent strips and the
Multistix Pro reagent strips (Siemens Medical
Solutions Diagnostics, Tarrytown, N.Y.) provide
simultaneous measurement of albu min/protein
and creatinine that permits an estimation of the
24-hr microalbumin excretion.3 As discussed in
Chapter 2, creatinine is produced and excreted at
a consistent rate for each individual. Therefore,
by comparing the albumin excre tion to the
creatinine excretion, the albumin reading can be
corrected for overhydration and dehydration in a
random sample. In additon to including creatinine
on the reagent strip, the albumin test pad is
changed to a dye-binding reac tion that is more
specific for albumin than the protein error of
indicators reaction on strips measuring protein.
Reagent Strip Reactions
Albumin
Albumin reagent strips utilize the dye bis(3′,3′′,
diodo-4′,4′′-
dihydroxy-5′,5′′-dinitrophenyl)-3,4,5,6-tetra-bromo
sulphonphtalein (DIDNTB), which has a higher
sensitivity and specificity for albumin. Whereas
conventional protein reagent pads have a
sensitivity ≥30 mg/dL and may include proteins
other than albumin, the DIDNTB strips can
measure albumin between 8 and 20 mg/dL (80 to
200 mg/L) without inclusion of other proteins.
Reaction interference by highly buffered alkaline
urine (always a concern with conventional
reagent strips) is controlled by using paper
treated with bis-(hep tapropylene glycol)
carbonate. Addition of polymethyl vinyl ether
decreases the nonspecific binding of polyamino
acids to the albumin pad. Results are reported as
10 to 150 mg/L (1 to 15 mg/dL). Colors range
from pale green to aqua blue. Falsely elevated
results can be caused by visibly bloody urine,
and abnormally colored urines may interfere with
the readings.7
Creatinine
The principle of the reagent strip for creatinine is
based on the pseudoperoxidase activity of
copper-creatinine complexes. The reaction
follows the same principle as the reaction for
blood on the reagent strips. Reagent strips
,
contain copper sul fate (CuSO4
3,3′,5,5′-tetramethylbenzidine (TMB) and diiso
propyl benzene dihydroperoxide (DBDH).
Creatinine in the
CHAPTER 5 • Chemical Examination of Urine 61
urine combines with the copper sulfate to form
copper creatinine peroxidase. This reacts with the
peroxide DBDH, releasing oxygen ions that oxidize
the chromogen TMB and producing a color change
from orange through green to blue.3
CuSO4 CRE → Cu(CRE) peroxidase
Cu(CRE) peroxidase
DBDH TMB → Oxidized TMB H2O
(peroxidase) (chromogen)
Results are reported as 10, 50, 100, 200, 300
mg/dL or 0.9, 4.4, 8.8, 17.7, or 26.5 mmol/L of
creatinine. Reagent strips are unable to detect the
absence of creati nine. Falsely elevated results can
be caused by visibly bloody urine and the presence
of the gastric acid–reducing medica tion cimetidine
(Tagamet). Abnormally colored urines also may
interfere with the readings.
No creatinine readings are considered
abnormal, as cre atinine is normally present in
concentrations of 10 to 300 mg/dL (0.9 to 26.5
mmol/L). The purpose of the creatinine
measurement is to correlate the albumin
concentraton to the urine concentration, producing
a semiquantitative albumin: creatinine ratio (A:C)
ratio.
Albumin/Protein:Creatinine Ratio
Automated and manual methods are available for
determining the A:C ratio based on the previously
discussed reactions. The Clinitek Microalbumin
reagent strips are designed for instru mental use
only. Strips are read on Clinitek Urine Chemistry
Analyzers. The strips measure only albumin and
creatinine and calculate the A:C ratio. Results are
displayed and printed out for albumin, creatinine,
and the A:C ratio in both con ventional and S.I.
units. Abnormal results for the A:C ratio are 30 to
300 mg/g or 3.4 to 33.9 mg/mmol.7
The Bayer Multistix Pro 11 reagent strips
include reagent pads for creatinine, protein-high
and protein-low (albumin), along with pads for
glucose, ketones, blood, nitrite, leukocyte esterase,
pH, bilirubin, and specific gravity. Urobilinogen is
not included on these strips. The strips can be read
manually or on automated Clinitek instruments.
The protein-high reac tion uses the protein error of
indicators principle and the pro tein-low reaction is
the previously discussed dye-binding method.
Results are reported as the protein:creatinine ratio,
although the protein-low result is also included in
the calcu lation. A manufacturer-supplied chart is
used to determine the ratio based on the results of
the protein-high, protein-low, and creatinine
readings.8
■ ■ ● Glucose
Because of its value in the detection and
monitoring of dia betes mellitus, the glucose test is
the most frequent chemical analysis performed on
urine. Owing to the nonspecific symp toms
associated with the onset of diabetes, it is
estimated that more than half of the cases in the
world are undiagnosed. Therefore, blood and urine
glucose tests are included in all physical
examinations and are often the focus of mass
health screening programs. Early diagnosis of
diabetes mellitus through blood and urine glucose
tests provides a greatly improved prognosis. Using
currently available reagent strip